Download (МО) in description of covalent and coordinate bonds.

Olga V. Kovalchukova
BASIC BIOINORGANIC CHEMISTRY
Educational and methodical complex
Moscow
Peoples’ Friendship University of Russia
2013
УДК
ББК
Б
Утверждено
РИС Ученого совета
Российского университета
дружбы народов
Издание подготовлено в рамках реализации
Программы стратегического развития РУДН на 2012–2016 гг.
Б
Ковальчукова, О. В.
Основы бионеорганической химии (Basic Bioinorganic
Chemistry) [Текст] : учебно-методический комплекс / О. В.
Ковальчукова. – М.: РУДН, 2013. – 214 с.
Представленный учебно-методический комплекс обеспечивает
реализацию магистерской программы «Химия окружающей среды» и
включает краткое описание курса, программу, систему оценки знаний,
краткий конспект лекций, описание основных лабораторных работ и
материалы и вопросы для подготовки к текущему и итоговому контролю
знаний.
The presented educational and methodical complex provides the
realization of the Master’s Program “Chemistry of the Environment”. It
includes the short description of the course, its syllabus, grading procedure,
lecture notes, description of the major lab. experiments, as well as the
materials and questions for the mid-semester and final controls.
ISBN 978-5-209-
© Ковальчукова О.В., 2013
© Российский университет дружбы народов, Издательство, 2013
2
INTRODUCTION
The course of the Basic Bioinorganic Chemistry is at the
gate-way of inorganic chemistry and biochemistry. It describes
the mutual relationship between these two sub-disciplines, with
focus upon the function of inorganic “substances“ in living
systems, including the transport, speciation and, eventually,
mineralisation of inorganic materials, and including the use of
inorganics in medicinal therapy and diagnosis. These
“substances” can be metal ions (such as Na+, Fe2+, Fe3+),
composite ions (e.g. MoO42-), coordination compounds (like
hemoglobin and cobalamine), or inorganic molecules (CO, NO,
O3).
The course begins with the principles of coordination
chemistry and a survey of the abilities of various functional
groups within proteins and nucleic acids to form coordination
complexes with metal ions.
The reactivity of coordination compounds of metal ions is
discussed in the context of the reaction mechanisms of specific
metalloenzymes. A part of the course is devoted to medicallyrelevant topics (e.g. the mechanisms by which organisms obtain
required metal ions from their environment, the toxicity of
metals, and use of metal containing compounds in treating cancer
and other deceases).
3
4
Content
Basic Bioinorganic Chemistry
(Study Guide)………………………………..
1.
2.
3.
4.
4.1.
4.2.
4.3.
5.
6.
7.
8.
9.
10.
Aims and Goals………………………………………
Requirements to results of development of the
discipline……………………………………………….
Filling of the discipline and study kinds……………
The discipline maintenance………………………….
The maintenance of sections of the discipline……….
The sections of the discipline and interdisciplinary
connections with the other disciplines……………….
The discipline maintenance and kinds of
employment…………………………………………….
Laboratory practical work……………………………
Subjects of research presentations……………………
Discipline information support………………………
Discipline Material support…………………………..
Methodical recommendations about the organization
of studying of the discipline…………………………..
Grading………………………………………………..
7
9
9
10
11
11
12
13
13
15
15
16
16
17
Supplementary Materials…………………….
19
Glossary of Terms…………………………….
21
Lecture notes………………………………….. 63
Chapter 1 Introduction to Bioinorganic Chemistry……..
Chapter 2 Transition Metal Chemistry………………….
65
88
5
Chapter 3 Structure, properties, and functions of metal
complexes of proteins and nucleic acids……...
Chapter 4 Transition Metal Storage, Transport and
Biomineralization………………………………
Chapter 5 Biological dioxygen transport system………...
Chapter 6 Bioinorganic Chemistry of dinitrogen
fixation…………………………………………..
107
126
138
161
Experiments in Bioinorganic Chemistry……. 169
6
Instructions for Research topic:
Presentation, Questions and Term Paper…
209
Exam Questions……………………………..
211
List of citations………………………….
213
7
BASIC BIOINORGANIC
CHEMISTRY
Study Guide
8
1. Aims and Goals
The aims and goals of the discipline are to provide
students in basic knowledges in properties of bio-essential
inorganic elements and their behavior in biological systems
The discipline is included in the variational part of the
Professional cycle. The subject maintenance is based on such
disciplines as “General Chemistry”, “Inorganic Chemistry”,
“Organic Chemistry”, “Analytical Chemistry”, and “Physical
Chemistry”.
2. Requirements to results of development of
the discipline
The teaching process is faced on formation of the
following competences:
ОК-1-6, ПК-1-7
As a result of studying of the discipline, students must:
Know: theoretical bases of bioinorganic chemistry,
composition and structure of complexes of metals with
constituents of biologically active molecules, principles of action
of metalloenzymes
Be able to: predict the electronic and spatial structures
of active sites of metalloenzymes; determine the composition and
stability of metallic complexes with the organic ligands – models
of biologically active molecules.
Own: the methods of investigation of biologically active
metallic complexes.
9
3. Filling of the discipline and study kinds
The total labor input includes 4 credits.
The study kinds
Auditoriums (in total)
Including:
Lectures
Practicals
Tutorials
Laboratory
Out-class work (in total)
Including:
Academic year project
Calculations
Research presentation
Slide presentation
Other types of work
(including interactive)
Final examination
Hours
72
I
72
36
36
36
72
36
72
8
20
20
20
4
The total labor input:
Hours
Credits
10
144
4
Semester
II
III
IV
4. The discipline maintenance
4.1. The maintenance of sections of the discipline
No
1.
The section
Introduction.
2.
Transitional metal
chemistry
3.
Structure, properties
and functions of
metal complexes of
proteins and nucleic
acids.
4.
Transition-Metal
Storage, Transport,
and
Biomineralization
5.
Biological dioxygen
The maintenance
Historical background and current
relevance. Bio-essential elements.
Fundamentals of Coordination
Chemistry.
Crystal Field Theory: Ligands &
Geometries: Oh; Td; Sq.Pl. 10Dq;
spectrochemical series; Template
effect; Chelate effect; Hard/Soft
Acids/Bases
Model Compounds
Composition of proteins and nucleic
acids. Aminoacids. Nucleic bases.
Electron donating groups and
features of their complex formation.
Chelating properties of aminoacids.
Complexes of transition metals with
proteins and nucleic acids and their
biological applications.
Biological significance of metals.
Chemical Properties Relative to
Storage and Transport. Role of
ferritin and transferring in storage
and transport of iron. Iron
biomineralization. Vitamins and Coferments.
Electronic structure and oxidative
11
transport system
6.
Bioinorganic
Chemistry of
dinitrogen fixation.
properties if dioxygen.
Requirements for Effective Oxygen
Carriers. Free-Radical Autoxidation.
Role of enzymes. Dioxygen
toxicity. Defense and Repair
Systems. Electron transfers in
biology.
Nitrogenase, Мо-Fе- and Feprotein. Dinitrogen complexes with
transitional metals. Chlorophyll as
Mg-comples, photosynthesis.
4.2. The sections of the discipline and
interdisciplinary connections with the other
disciplines
No
Name of the connecting
discipline
1.
Chemistry of the
enviroments and
ecological problems
Instrumental methods
of analysis of natural
compounds
Physical and chemical
methods of studying of
chemical compounds
Chemical toxicology
Philosophical problems
of chemistry
2.
3.
4.
5.
12
No of the section if the studying
discipline
1
2
3
4
5
6
+
+
+
+
+
+
+
+
4.3. The discipline maintenance and kinds of
employment
No
1.
2.
3.
4.
5.
6.
The name of the
section of the
discipline
Introduction.
Transitional metal
chemistry
Structure, properties
and functions of
metal complexes of
proteins and nucleic
acids.
Transition-Metal
Storage, Transport,
and
Biomineralization
Biological dioxygen
transport system
Bioinorganic
Chemistry of
dinitrogen fixation.
OutLect Pract Lab Tut class
work
6
6
6
4
6
In
total,
hours
12
16
6
10
26
6
8
10
24
6
4
6
16
6
4
4
14
10
5. Laboratory practical work
No
No of the
Name of the laboratory practical
section of the
work
discipline
1.
1
Electronic formulae of atoms and
ions. Molecular diagrams of
labor
input
(hours)
2
13
2.
1
3.
2
4.
3
5.
3
6.
4
7.
3
8.
4
9.
5, 6
10.
1–6
14
molecules and molecular ions.
Bond orders and multiplicities of
electronic states.
Electronic and spatial structures
and chemical properties of
coordinate compounds.
Redox-properties of complexes of
transition metals.
Quantum-chemical modeling of
tautomeric forms of nucleic acids
and their complexes with metals.
Determination of ionization
constants of amino acids by
potentiometry.
Determination of stability
constants of complexes of
transitional metals by
potentiometry.
Determination of protonation
constants of nucleic bases by UVspectrophotometry.
Determination of stability
constants of complexes of
transitional metals with the nuclear
bases by UV-spectrophotometry.
Determination of coordination
sites of O- and N-donating ligands
by IR-spectroscopy.
Conference on research topics.
2
2
4
4
4
4
4
6
4
6. Subjects of research presentations
1.
2.
3.
4.
5.
6.
7.
8.
9.
10.
11.
X-Ray and DNA demonstration.
Storage, Transport, and Biomineralization of Zink.
Storage, Transport, and Biomineralization of Copper.
Storage, Transport, and Biomineralization of Cobalt.
Molybdenum enzymes.
Selective organic synthesis catalyzed by organometallic
compounds.
Mechanisms of metallic toxicity.
Chelation therapy and metallotherapy.
Manganese complexes as Catalysts for Peroxynitrite
Decomposition.
Oxidative stress and its influence on organisms.
Metals and Health.
7. Discipline information support
Required Textbook:
Ivano Bertini, Harry B. Gray,, Bioinorganic Chemistry, Mill
Valley, California, 1994. (available at the Department of General
Chemistry)
Supplementary readings:
i) S.J. Lippard, J.M. Berg, Principles of Bioinorganic
Chemistry, U.S.B., 1994.
ii) Frausto da Silva, J. J. R. and Williams, R. J. P. The
Biological Chemistry of the Elements: the Inorganic
Chemistry of Life (2nd. edition, Oxford, 2001).
iii) Roat-Malone, R. M. Bioinorganic Chemistry: A Short
Course (Wiley, 2002).
15
iv) Bertini, I.; Gray, H. B.; Stiefel, E. I.; Valentine, J. S.,
editors. Biological Inorganic Chemistry (University
Science Books, 2007).
v) Journal of Inorganic Biochemistry (original papers).
Computational support:
AM1, PM3
Databases, directory and search engines:





http://www.sciencedirect.com/science/journal/00063061
http://www.chm.bris.ac.uk/pt/harvey/bioinorganic.htm
http://wwwchem.uwimona.edu.jm:1104/courses/CFT.html
http://www.pnas.org/content/100/7/3601.full
http://books.google.ru/books?id=zGJtXzPINAUC&printsec=f
rontcover&hl=ru#v=onepage&q&f=false
 http://www.adichemistry.com/inorganic/bioinorganic/bioinor
ganic-chemistry.html
 http://simple.wikipedia.org/wiki/
8. Discipline Material support
Chemical
equipment
and
reagents,
spectrophotometers, pH-meter, computers
IR
and
UV
9. Methodical recommendations about the
organization of studying of the discipline
During studying the discipline the students attend lectures,
perform lab experiments and prepare a selected research topic for
16
presentation and discussion in the class or in the interactive form.
Each kind of study is graded by special No of points.
10. Grading
The study kind
1. Lecture attendance
2. Laboratory experiments
3. Research topic
In total
No of points
95 - 100
86 - 94
69 - 85
61 - 68
51 - 60
31 - 50
0 - 30
No of
kinds
18
10
1
Grade
5+
5
4
3+
3
2+
2
No of
points
0-2
0-5
0-14
Maximal
points
36
50
14
100
ECTS grade
A
B
C
D
E
FX
F
17
18
SUPPLEMENTARY MATERIALS
19
20
GLOSSARY OF TERMS1
1
Cited from Glossary of terms used in Bioinorganic Chemistry.
Ed. M. W. G. de Bolster. Pure &App. Chem., Vol. 69, No. 6, pp.
1251-1303, 1997.
21
22
Acid
See Brensted acid, Lewis acid, hard acid,
soft acid.
Acidity constant
The equilibrium constant for splitting off a
hydron from a Brensted acid.
Active center
The location in an enzyme where the
specific reaction takes place.
Active site
See active center.
Aerobe
An organism that needs dioxygen for
respiration and thus for growth.
Albumin
A type of protein, especially a protein of
blood plasma which transports various
substances, including metal ions, drugs and
xenobiotics.
Ambidentate
Ligands, such as (NCS)-, that can bind to a
central atom through either of two or more
donor atoms are termed ambidentate.
Amino-acid
residue (in a
polypeptide)
When two or more amino acids combine to
form a peptide, the elements of water are
removed, and what remains of each amino
acid is called an amino-acid residue. Aminoacid residues are therefore structures that
lack a hydrogen atom of the amino group
(-NH-CHR-COOH), or the hydroxy moiety
of the carboxy group (NH,-CHR-CO-), or
both
(-NH-CHR-CO-); all units of a
23
peptide chain are therefore amino-acid
residues. (Residues of amino acids that
contain two amino groups or two carboxy
groups may be joined by isopeptide bonds,
and so may not have the formulas shown).
The residue in a peptide that has an amino
group that is free, or at least not acylated by
another amino-acid residue (it may, for
example, be acylated or formylated), is
called N-terminal; it is the N-terminus. The
residue that has a free carboxy group, or at
least does not acylate another amino-acid
residue, (it may, for example, acylate
ammonia to give -NHCHR-CO-NH,), is
called C-terminal.
Anabolism
The processes of metabolism that result in the
synthesis of cellular components from
precursors of low molecular weight.
Anaerobe
An organism that does not need dioxygen
for growth. Many anaerobes are even
sensitive to dioxygen. Obligate (strict)
anaerobes grow only in the absence of
dioxygen. Facultative anaerobes can grow
either in the presence or in the absence of
dioxygen.
Anation
Replacement of the ligand water by an anion
in a coordination entity.
Anti
In the representation of stereochemical
relationships “anti” means “on opposite
24
sides” of a reference plane, in contrast to
“syn” which means “on the same side”.
Aquation
The incorporation of one or more integral
molecules of water into another chemical
species with or without displacement of one
or more other atoms or groups. See also
hydration.
ATP
adenosine 5’-triphosphate.
Auranofin
See gold drugs.
Azurin
An electron transfer protein containing a
type 1 copper site, isolated from certain
bacteria.
Base
See Bronsted base, Lewis base, hard base,
soft base.
Base pairing
The specific association between two
complementary strands of nucleic acids that
results from theformation of hydrogen bonds
between the base components of the
nucleotides of each strand: A=T and G=C in
DNA, A=U and Gr-C (and in some cases
Gr-U) in RlVA (the lines indicate the
number of hydrogen bonds). Single-stranded
nucleic acid molecules can adopt a partially
double-stranded structure throughintrastrand
base pairing. See also nucleosides.
Basicity constant
See acidity constant.
25
Beta() sheet
Preferentially called a beta pleated sheet; a
regular structure in an extended polypeptide
chain, stabilized inthe form of a sheet by
hydrogen bonds between CO and NH groups
of adjacent (parallel or antiparallel)chains.
Beta() strand
Element of a beta sheet. One of the strands
that is hydrogen bonded to a parallel or
antiparallel strand to form a beta sheet.
Beta() turn
A hairpin structure in a polypeptide chain
reversing its direction by forming a
hydrogen bond between the CO group of
amino-acid residue n with the NH group of
residue (n+3).
Bifunctional
ligand
A ligand that is capable of simultaneous use
of two of its donor atoms to bind to one or
more central atoms (see also ambidentate).
Binding constant
See stability constant.
Binding site
A specific region (or atom) in a molecular
entity that is capable of entering into a
stabilizing interaction with another
molecular entity. An example of such an
interaction is that of an active site in an
enzyme with its substrate. Typical forms of
interaction are by hydrogen bonding,
coordination, and ion pair formation. Two
binding sites in different molecular entities
are said to be complementary if their
26
interaction is stabilizing.
Binuclear
Less-frequently used term for the IUPAC
recommended: dinuclear. See nuclearit.y
Biocatalyst
A catalyst of biological origin, typically an
enzyme.
Bioleaching
Extraction of metals from ores or soil by
biological processes, mostly by
microorganisms.
Biomembrane
Organized sheet-like assemblies consisting
mainly of proteins and lipids (bilayers),
acting as highly selective permeability
barriers, containing specific molecular
pumps and gates, receptors and enzymes.
Biomimetic
Refers to a laboratory procedure designed to
imitate a natural chemical process. Also
refers to a compound that mimics a
biological material in its structure or
function.
Biomineralization The synthesis of inorganic crystalline or
amorphous mineral-like materials by living
organisms. Among the minerals synthesized
biologically in various forms of life are:
fluoroapatite, hydroxyapatite (Ca5(PO4)3(F,
OH)), magnetite (Fe3O4) and calcium
carbonate (CaCO3).
Biopolymers
Macromolecules (including proteins, nucleic
27
acids and polysaccharides) formed by living
organisms.
Biotransformatio
n
A chemical transformation mediated by
living organisms or enzyme preparations.
See also bioconversion.
Blue copper
protein
An electron transfer protein containing a
type 1 copper site. Characterized by a strong
absorption in the visible region, and an EPR
signal with an unusually small hyperfine
coupling to the copper nucleus.
Bridging ligand
A bridging ligand binds to two or more
central atoms, usually metals, thereby
linking them together to produce
polynuclear coordination entities . Bridging
is indicated by the Greek letter p appearing
before the ligand name and separated by a
hyphen. For an example, see FeMocofactor.
Brønsted acid
A molecular entity capable of donating a
hydron to a base (i.e., a “hydron donor”) or
the corresponding chemical species.
Brønsted base
A molecular entity capable of accepting a
hydron from an acid (i.e., a “hydron
acceptor”) or the corresponding chemical
species.
Carbon monoxide Enzymes that catalyze the oxidation of
carbon monoxide to carbon dioxide. They
dehydrogenases
28
contain iron-sulfur clusters and either
nickel and zinc, or molybdopterin. Some
nickel-containing enzymes are also involved
in the synthesis of acetyl coenzyme A from
CO2 and H2.
Catalase
A hemeprotein, which catalyzes the
disproportionation of dihydrogen peroxide
to O2 and water ; it also catalyzes the
oxidation of other compounds, such as
ethanol, by dihydrogen peroxide. A nonheme protein containing a dinuclear
manganese cluster with catalase activity is
often called pseudocatalase.
Central atom
The atom in a coordination entity that binds
other atoms or group of atoms (ligands) to
itself, thereby occupying a central position
in the coordination entity.
Charge-transfer
complex
An aggregate of two or more molecules in
which charge is transferred from a donor to
an acceptor.
Chelation
Chelation involves coordination of more
than one sigma-electron pair donor group
from the same ligand to the same central
atom. The number of coordinating groups
in a single chelating ligand is indicated by
the adjectives didentate, tridentate,
tetradentate, etc.
Chlorophyll
Part of the photosynthetic systems in green
29
plants. Generally speaking, it can be
considered as a magnesium complex of
aporphyrin in which a double bond in one
of the pyrrole rings (17-18) has been
reduced. A fused cyclopentanone ring is also
present (positions 13-14-15). In the case of
chlorophyll a, the substituted porphyrin
ligand further contains four methyl groups in
positions 2,7, 12 and 18, a vinyl group in
position 3, an ethyl group in position 8 and
a -(CH,),CO,R group (R = phytyl, (2-(7R,
11R)-3,7,11,15-tetramethylhexadec-2-en-1yli)n position 17. In chlorophyll b, the group
in position 7 is a -CHO group. In
bacteriochlorophyll a the porphyrin ring is
further reduced (7-8), and the group in
position 3 is now a -COCH, group. In
addition, in bacteriochlorophyll b, the group
in position 8 is a = CHCH, group.
30
Cluster
A number of metal centers grouped close
together which can have direct metal
bonding interactions or interactions through
a bridging ligand, but are not necessarily
held together by these interactions.
Examples can be found under the entries
r2Fe-25'1, r4Fe-45'1, ferredoxin, HiPIP,
ironaulfur cluster, FeMo-cofactor, ferritin,
metallo- thionein, nitrogenase, and Rieske
iron-suwur protein.
Cobalamin
Vitamin B-12 or B12. A vitamin synthesized
by microorganisms and conserved in
animals in the liver. Deficiency or collective
uptake of vitamin B-12 leads to pernicious
anemia. Cobalamin is a substituted corrinCo(III) complex in which the cobalt atom is
bound to the four nitrogen atoms of the
corrin ring, an axial group R and 5,6dimethylbenzimidazole.The latter is linked
to the cobalt by the N-3 nitrogen atom and is
bound to the C-1 carbon of a ribose
molecule by the N-1 nitrogen atom. Various
forms of the vitamin are known with
different R groups such as R = CN,
cyanocobalamin; R = OH,
hydroxocobalamin; R = CH3
methylcobalamin; R = 5'-deoxyadenosyl,
coenzyme B-12.
Coenzyme
A low-molecular-weight, non-protein
organic compound (often a nucleotide)
31
participating in enzymatic reactions as
dissociable acceptor or donor of chemical
groups or electrons.
Cofactor
An organic molecule or ion (usually a metal
ion) that is required by an enzyme for its
activity. It may be attached either loosely
(coenzyme) or tightly (prosthetic group).
Configuration
In the context of stereochemistry, the term is
restricted to the arrangements of atoms of a
molecular entity in space that distinguishes
stereoisomers, the isomerism between which
is not due to conformational differences.
Conformation
The spacial arrangements of atoms affording
distinction between stereoisomers which can
be interconverted by rotations about
formally single bonds.
Cooperativity
The phenomenon that binding of an effector
molecule to a biological system either
enhances or diminishes the binding of a
successive molecule, of the same or different
kind, to the same system. The system may
be an entyme, or a protein that specifically
binds another molecule such as oxygen or
DNA. The effector molecule may be an
enzyme substrate or an allosteric effector.
The enzyme or protein exists in different
conformations, with different catalytic rates
or binding affinities, and binding of the
effector molecule changes the proportion of
32
these conformations. Enhanced binding is
named positive cooperativity; diminished
binding is named negative cooperativity. A
well-known example of positive
cooperativity is in hemoglobin, In
biocatalysis it was originally proposed that
only multi-subunit enzymes could respond
in this way. However single-subunit
enzymes may give such a response (socalled mnemonic enzymes).
Coordination
A coordination entity is composed of a
central atom, usually that of a metal, to
which is attached a surrounding array of
other atoms or group of atoms, each of
which is called a ligand. A coordination
entity may be a neutral molecole, a cation or
an anion. The ligands may be viewed as
neutral or ionic entities that are bonded to an
appropriately charged central atom. It is
standard practice to think of the ligand
atoms that are directly attached to the central
atom as defining a coordination polyhedron
(tetrahedron, square plane, octahedron, etc.)
about the central atom. The coordination
number is defined as being equal to the
number of sigma-bonds between ligands and
the central atom; this definition is not
necessarily appropriate in all areas of
(coordination) chemistry. In a coordination
formula, the central atom is listed first. The
formally anionic ligands appear next and
they are listed in alphabetic order according
33
to the first symbols of their formula. The
neutral ligands follow, also in alphabetical
order, according to the same principle. The
formula for the entire coordination entity,
whether charged or not, is enclosed in
square brackets. In a coordination name, the
ligands are listed in alphabetical order,
without regard to charge, before the name of
the central atom. Numerical prefixes
indicating the number of ligands are not
considered in determining that order. All
anionic coordination entities take the ending
-ate, whereas no distinguishing termination
is used for cationic or neutral coordination
entities.
Crystal field
34
Crystal field theory is the theory which
interprets the properties of coordination
entities on the basis that the interaction of
the ligands and the central atom is a strictly
ionic or ion-dipole interaction resulting from
electrostatic attractions between the central
atom and the ligands. The ligands are
regarded as point negative (or partially
negative) charges surrounding a central
atom; covalent bonding is completely
neglected. The splitting or separation of
energy levels of the five degenerate d
orbitals in a transition metal, when the metal
is surrounded by ligands arranged in a
particular geometry with respect to the metal
center, is called the crystal field splitting.
See also ligandjield.
C-terminal
amino-acid
residue
See amino-acid residue.
Cytochrome
A heme protein that transfers electrons, and
exhibits intense absorption bands (between
510 and 615 nm in the reduced form.
Cytochromes are designated types a, b, c or
d, depending on the type of heme. The iron
undergoes oxidation-reduction between
oxidation states Fe(II) and Fe(III). Most
cytochromes are hemochromes, in which the
fifth and sixth coordination sites in the iron
are occupied by strong field ligands,
regardless of the oxidation state of iron.
Cytochrome-c
oxidase
An eniyme, ferrocytochrome-c: dioxygen
oxidoreductase. The major respiratory
protein of animal and plant mitochondria, it
catalyzes the oxidation of Fe(II)-cytochrome
c, and the reduction of dioxygen to water.
Contains two hemes and three copper atoms,
arranged in three centers. Heme and copper
form a center that reacts with dioxygen; the
second heme is cytochrome; the third site,
copper, is a dinuclear center.
Cytochrome P450
General term for a group of heme-containing
monooxygenases . Named from the
prominent absorption band of the Fe(II)carbonyl complex. The heme comprises
protophorphyrin, and the proximal ligand to
35
iron is a cysteine sulfur. Cytochromes P-450
of microsomes in tissues such as liver are
responsible for metabolism of many
xenobiotics, including drugs. Others, such as
the mitochondrial eniymes from adrenal
glands, are involved in biosynthetic
pathways such as those of steroids. The
reaction with dioxygen appears to involve
higher oxidation states of iron, such as
Fe(IV)=O.
Dehydrogenase
An oxidoreductase which catalyzes the
removal of hydrogen atoms from a substrate.
Denitrification
The reduction of nitrates to nitrites, nitrogen
monoxide (nitric oxide), dinitrogen oxide
(nitrous oxide) and ultimately dinitrogen
catalyzed by microorganisms, e.g.
facultative aerobic soil bacteria under
anaerobic conditions.
Denticity
The number of donor groups from a given
ligand attached to the same central atom.
Deoxyribonucleic A high-molecular-mass linear polymer
composed of nucleotides containing 2acid (DNA)
deoxyribose and linked between positions 3'
and 5' by phosphodiester groups; DNA
contains the genetic information of
organisms. The double-stranded form
consists of a double helix of two
complementary chains that run in opposite
directions and are held together by hydrogen
36
bonds between pairs of the complementary
nucleotides. The way the helices are
constructed may differ and is usually
designated as A, B, Z, etc. Occasionally,
alternative structures are found, such as
those with Hoogsteen base pairing.
Desferrioxamine
(dfo)
Chelating agent used world-wide in the
treatment of iron overload conditions, such
as hemochromatosis and thalassemia.
Diamagnetic
Substances having a negative magnetic
susceptibility are diamagnetic. They are
repelled by a magnetic field.
Dinuclear
See nuclearity.
Dioxygenase
An enzyme that catalyzes the insertion of
two oxygen atoms into a substrate, both
oxygens being derived from O2.
Dismutase
An enzyme that catalyzes a
disproportionation reaction.
Dismutation
See disproportionation.
Disproportionati
on
Any chemical reaction of the type A + A +
A' + A" where A, A' and A'' are different
chemical species. The reverse of
disproportionation (or dismutation) is called
comproportionation.
Dissociation
See stability constant.
37
constant
DNA
See deoxyribonucleic acid.
Domain
An independently folded unit within a
protein, often joined by a flexible segment
of the polypeptide chain.
Donor atom
symbol
A polydentate ligand possesses more than
one donor site, some or all of which may be
involved in coordination. To indicate the
points of ligation, a system is needed. The
general and systematic system for doing this
is called the kappa convention: single
ligating atom attachments of a polyatomic
ligand to a coordination centre are indicated
by the italic element symbol preceded by a
Greek kappa, . In earlier practice, the
different donors of the ligand were denoted
by adding to the end of the name of the
ligand the italicized symbol(s) for the atom
or atoms through which attachment to the
metal occurs.
Electron transfer
protein
A protein, often containing a metal ion that
oxidizes and reduces other molecules by
means of electron transfer.
Endogenous
Originating internally. In the description of
metal ion coordination in metalloproteins,
endogenous refers to internal, or proteinderived, ligands.
38
Entatic state
A state of an atom or group which due to its
binding in a protein has its geometric or
electronic condition adapted for function.
Enzyme
A macromolecule that functions as a
biocatalyst by increasing the reaction rate,
frequently containing or requiring one or
more metal ions. In general, an enzyme
catalyzes only one reaction type (reaction
specificity) and operates on only a narrow
range of substrates (substrate specificity).
Substrate molecules are attacked at the same
site (regiospecificity) and only one or
preferentially one of the enantiomers of
chiral substrate or of racemic mixtures is
attacked (enantiospecificity).
Equilibrium
constant
See acidity constant and stability constant.
Exogenous
Originating externally. In the context of
metalloprotein ligands, exogenous describes
ligands added from an external source, such
as CO or O2.
FeMo-cofactor
An inorganic cluster that is found in the
FeMo protein of the molybdenumnitrogenae and is essential for the catalytic
reduction of N, to ammonia. This cluster
contains Fe, Mo and S in a 7: 1 : 9 ratio. The
structure of the cofactor within the FeMo
protein can be described in terms of two
cuboidal subunits, Fe,S, and MoFe,S,
39
bridged by three S2- ions and “anchored” to
the protein by a histidine bound via an
imidazole group to the Mo atom and by a
cysteine bound via a deprotonated SH group
to an Fe atom of the Fe,S, subunit. The Mo
atom at the periphery of the molecule is sixcoordinate and in addition to the three
sulfide-ligands and the histidine imidazole is
also bound to two oxygen atoms from an
(R)-homocitrate molecule.
Fenton reaction
Fe2+ + H2O2  Fe3+ + OH + OH-. This is
the iron-salt-dependent decomposition of
dihydrogen peroxide, generating the highly
reactive hydroxyl radical, possibly via an
oxoiron(1V) intermediate. Addition of a
reducing agent such as ascorbate leads to a
cycle which increases the damage to
biological molecules. See also Haber-Weiss
reaction.
Ferredoxin
A protein containing more than one iron and
acid-labile sulfur, that displays electrontransfer activity but not classical enzyme
function. See also HiPIP.
40
Ferriheme
An iron(III) porphyrin coordination
complex.
Ferritin
An iron storage protein consisting of a shell
of 24 protein subunits, encapsulating up to
4500 iron atoms in the form of a hydrated
iron(III) oxide.
Ferrochelatase
An enzyme which contains an Fe-S cluster
and catalyzes the insertion of iron into
protoporphyrin to form heme..
Ferroheme
An iron(II) porphyrin coordination
complex.
Formation
constant
See stability constant.
Gold drugs
Gold coordination compounds used in the
treatment of rheumatoid arthritis, examples
being auranofin, (tetraacetylthioglucosatoS)(triethylphosphane)gold(I), and
myocrysin, disodium thiomalatogold(I).
Haber-Weiss
reaction
The Haber-Weiss cycle consists of the
following two reactions :
H2O2 + OH  H2O + O2- + H+ and
H2O2 + O2  O2 + OH + OHThe second reaction achieved notoriety as a
possible source of hydroxyl radicals.
However, it has a negligible rate constant. It
is believed that iron(II1) complexes can
41
catalyze this reaction: first Fe(II1) is reduced
by superoxide, followed by oxidation by
dihydrogen peroxide. See also Fenton
reaction.
Hard acid
A Lewis acid with an acceptor centre of low
polarizability. It preferentially associates
with hard bases rather than with soft bases,
in a qualitative sense (sometimes called
“HSAB rule”). Conversely a soft acid
possesses an acceptor centre of high
polarizability and exhibits the reverse
preference for a partner for coordination.
Hard base
A Lewis base with a donor centre of low
polarizability; the converse applies to soft
bases. See also hard acid.
Heme
A near-planar coordination complex
obtained from iron and the dianionic form of
porphyrin. Derivatives are known with
substituents at various positions on the ring
named a, b, c, d, etc. Heme b, derived from
protoporphyrin, is the most frequently
occurring heme.
Hemocyanin
A dioxygen-carrying protein (from
invertebrates, e.g. arthropods and molluscs),
containing dinuclear type 3 copper sites.
Hemoglobin
A dioxygen-carrying heme protein of red
blood cells, generally consisting of two
alpha and two beta subunits, each
42
containing one molecule of protoporphyrin.
High-spin
See low-spin.
HiPlP
Formerly-used abbreviation for HighPotential Iron-sulfur Protein, now classed
as a ferredoxin. An electron transfer
protein from photosynthetic and other
bacteria, containing a [4Fe-4S] cluster
which undergoes oxidation-reduction
between the [4Fe-4S]2+ and [4Fe-4S]3+
states.
Hydration
Addition of water or the elements of water
(i.e. H and OH) to a molecular entity. The
term is also used in a more restricted sense
for the process: A (gas)  A (aqueous
solution); cf. the use of the term in inorganic
and physical chemistry to describe the state
of ions of an aqueous electrolyte solution.
See also aquation and salvation.
Hydrogenase
An envme, dihydrogen:acceptor
oxidoreductase, that catalyzes the formation
or oxidation of H2. Hydrogenases are of
various types. One class ([Fe]-hydrogenases)
contains only iron-sulfur clusters. The other
major class ([NiFe]-hydrogenases) has a
nickel-containing center and iron-sulk
clusters; a variation of the latter type
([NiFeSe]-hydrogenases) contains
selenocysteine.
43
Hydrolase
An enqvme of EC class 3, also known as a
hydro-lyase, that catalyzes the hydrolysis of
a substrate.
Hydrolysis
Solvolysis by water.
Hydrophilic
The term is used to mean "water preferring".
May also be used to describe the character
of interaction of a particular atomic group
with the medium.
Hydrophobic
interaction
The tendency of hydrocarbons (or of
lipophilic hydrocarbon-like groups in
solutes) to form intermolecular aggregates in
an aqueous medium, and analogous
intramolecular interactions. The name arises
from the attribution of the phenomenon to
the apparent repulsion between water and
hydrocarbons. Use of the misleading
alternative term hydrophobic bond is
discouraged.
Inhibition
The decrease in the rate of a reaction
brought about by the addition of a substance
(inhibitor).
Intercalation
compounds
Compounds resulting from inclusion,
usually without covalent bonding, of one
kind of molecule (the guest molecule) in a
matrix of another compound (the host
compound), which has a layered structure.
The host compound, with a rather rigid
structure, may be macromolecular,
44
crystalline, or amorphous.
Iron-responsive
element
A specific base sequence in certain
messenger RNAs that code for various
proteins of iron metabolism, which allows
regulation at translational level by the ironreponsive protein.
Iron-responsive
protein (IRP)
A protein that responds to the level of iron in
the cell, and regulates the biosynthesis of
proteins of iron metabolism, by binding to
the iron-responsive element on messenger
RNA.
Iron-sulfur
proteins
Proteins in which non-heme iron is
coordinated with cysteine sulfur and usually
also with inorganic sulfur. Divided into three
major categories: rubredoxins; "simple ironsulfur proteins", containing only ironsurfur
clusters, and "complex iron-sulfur proteins",
containing additional active redox centers
such as flavin, molybdenum or heme. In
most iron-sulfur proteins the clusters
function as electron transfer groups, but in
others they have other functions such as
catalysis of hydrataseldehydratase reactions,
maintenance of protein structure, or
regulation of activity.
lsomerase
An enzyme of EC class 5, which catalyzes
the isomerization of a substrate.
Labile
The term has loosely been used to describe
45
either a relatively unstable and transient
chemical species or a relatively stable but
reactive species.
Lewis acid
A molecular entity that is an electron pair
acceptor and therefore able to react with a
Lewis base to form a Lewis adduct, by
sharing the electron pair furnished by the
Lewis base.
Lewis adduct
The adduct formed between a Lewis acid
and a Lewis base.
Lewis base
A molecular entity able to provide a pair of
electrons and thus capable of coordination
to a Lewis acid, thereby producing a Lewis
adduct.
Ligand
In inorganic chemistry the ligands are the
atoms or groups of atoms bound to the
central atom (see also coordination). The
root of the word is sometimes converted into
the verb to ligate, meaning to coordinate as a
ligand, and the derived participles, ligating
and ligated. This use should not be confused
with its use to describe the action of ligases
(a class of enzymes). The names for anionic
ligands, whether inorganic or organic, end in
-O. In general, if the anion name ends in ide, or -ate, the final -e is replaced by -O,
giving ido, and -ato, respectively. Neutral
and cationic ligand names are used without
modification. Ligands bonded by a single
46
carbon atom to metals are regarded as
radical substituents, their names being
derived from the parent hydrocarbon, from
which one hydrogen atom has been
removed. In general, the final letter -e of the
name is replaced by -yl. In biochemistry the
term ligand has been used more widely: if it
is possible or convenient to regard part of a
polyatomic molecular entity as central, then
the atoms or groups or molecules bound to
that part may be called ligands.
Ligand field
Ligand field theory is a modified crystal
field theory that assigns certain parameters
as variables rather than taking them as equal
to the values found for free ions, thereby
taking into account the potential covalent
character of the metal-ligand bond.
Low-spin
In any coordination entity with a particular
dn (1 < n < 9) configuration and a particular
geometry, if the n electrons are distributed
so that they occupy the lowest possible
energy levels, the entity is a low-spin
complex. If some of the higher energy d
orbitals are occupied before all the lower
energy ones are completely filled, then the
entity is a high-spin complex.
Metabolism
The entire physical and chemical processes
involved in the maintenance and
reproduction of life in which nutrients are
broken down to generate energy and to give
47
simpler molecules (catabolism) which by
themselves may be used to form more
complex molecules (anabolism).
Metalloenzyme
An enzyme that, in the active state, contains
one or more metal ions which are essential
for its biological function.
Mixed valency
This is one of several names, such as ‘mixed
oxidation state’ or ‘non-integral oxidation
state’ used to describe coordination
compounds and clusters in which a metal is
present in more than one level of oxidation.
The importance in biology is due to the
often complete delocalization of the valence
electrons over the cluster, allowing efficient
electron-transfer processes. See also
oxidation number.
Model
A synthetic coordination entity that closely
approaches the properties of a metal ion in a
protein and yields useful information
concerning biological structure and function.
Given the fact that the term is also loosely
used to describe various types of molecular
structures, constructed, for example, in the
computer, the term biomimetic is more
appropriate.
Monooxygenase
An enzyme that catalyzes the insertion of
one atom of oxygen, derived from O,, into
an aromatic or aliphatic compound. The
reaction is coupled to the oxidation of a co-
48
substrate such as NAD(P)H or 2oxoglutarate.
Motif
A pattern of amino acids in a protein
sequence which has a specific function, e.g.
metal binding. See also consensus
sequence.
Multicopper
oxidases
A group of enzymes that oxidize organic
substrates and reduce dioxygen to water.
These contain a combination of copper ions
with different spectral features; called type I
centers, type 2 centers, and type 3 centers,
where the type 2 and type 3 sites are
clustered together as a trinuclear unit. Wellknown examples are: laccase, ascorbate
oxidase and ceruloplasmin.
Myocrysin
See gold drugs.
Myoglobin
A monomeric dioxygen-binding heme
protein of muscle tissue, structurally similar
to a subunit of hemoglobin.
Nitrate reductase
A metalloenzyme, containing molybdenum
that reduces nitrate to nitrite.
Nitrite reductase
A metalloenzyme that reduces nitrite.
Dissimilatory nitrite reductases contain
copper and reduce nitrite to nitrogen
monoxide. Assimilatory nitrite reductases
contain siroheme and iron-sulfur clusters
and reduce nitrite to ammonia.
49
Nitrogenase
An enzyme complex (EC 1.18.6.1) from
bacteria that catalyzes the reduction of
dinitrogen to ammonia:
N2 + 8e- + 10H+  2NH4+ + H2
with the simultaneous hydrolysis of at least
16 ATP molecules. The electron donor is
reduced ferredoxin or flavodoxin.
Dihydrogen is always a co-product of the
reaction. Ethyne (acetylene) can also be
reduced to ethene (ethylene) and in some
cases ethane. All nitrogenases are ironsulfur proteins. Three different types which
differ in the type of cofactor present have
been identified: molybdenum-nitrogenase
(the most common, which contains the ironmolybdenum cofactor), vanadiumnitrogenase, and iron-only nitrogenase. See
also FeMo-cofactor.
Nitrogen fixation
The assimilation of dinitrogen by microbial
reduction to ammonia and conversion into
organonitrogen compounds such as amino
acids. Only a limited number of
microorganisms are able to fix nitrogen. See
also nitrogenase.
N-terminal
amino-acid
residue
See amino-acid residue.
Nuclearity
The number of central atoms joined in a
single coordination entity by bridging
50
ligands or metal-metal bonds is indicated by
dinuclear, trinuclear, tetranuclear,
polynuclear, etc.
Nucleic acids
Macromolecules composed of sequences of
nucleotides that perform several functions in
living cells, e.g. the storage of genetic
information and its transfer from one
generation to the next (DNA), and the
expression of this information in protein
synthesis (mRNA, tRNA), and may act as
functional components of subcellular units
such as ribosomes (rRNA). RNA contains
D-ribose, DNA contains 2-deoxy-D-ribose
as the sugar component. Currently, synthetic
nucleic acids can be made consisting of
hundreds of nucleotides. See also
oligonucleotide.
Nucleosides
Compounds in which a purine or pyrimidine
base is -N-glycosidically bound to C-1 of
either 2-deoxy-D-ribose or of D-ribose, but
without any phosphate groups. The common
nucleosides in biological systems are
adenosine, guanosine, cytidine, and uridine
(which contain ribose) and deoxyadenosine,
deoxyguanosine, deoxycytidine and
thymidine (which contain deoxyribose).
Nucleotides
Nucleosides with one or more phosphate
groups esterified mainly to the 3’- or the 5’position of the sugar moiety. Nucleotides
found in cells are adenylic acid, guanylic
51
acid, uridylic acid, cytidylic acid,
deoxyadenylic acid, deoxyguanylic acid,
deoxycytidylic acid and thymidylic acid. See
also adenosine 5 triphosphate, NAD+,
NADP.
Oxidase
An enzyme which catalyzes the oxidation of
substrates by O2.
Oxidation
number
The oxidation number of an element in any
chemical entity is the number of charges
which would remain on a given atom if the
pairs of electrons in each bond to that atom
were assigned to the more electronegative
member of the bond pair. The oxidation
(Stock) number of an element is indicated by
a roman numeral placed in parentheses
immediately following the name (modified
if necessary by an appropriate ending) of the
element to which it refers. The oxidation
number may be positive, negative or zero.
Zero, not a roman numeral, is represented by
the usual cipher, 0. The positive sign is
never used. An oxidation number is always
positive unless the minus sign is explicitly
used. Note that it cannot be nonintegral (see
also mixed valency). Non-integral numbers
may seem appropriate in some cases where a
charge is spread over more than one atom,
but such a use is not encouraged. In such
ambiguous cases, the charge number, which
designates ionic charge, can be used. A
charge (Ewens-Bassett) number is a number
52
in parentheses written without a space
immediately after the name of an ion, and
whose magnitude is the ionic charge. Thus
the number may refer to cations or anions,
but never to neutral species. The charge is
written in arabic numerals and followed by
the sign of the charge. In a coordination
entity, the oxidation number of the central
atom is defined as the charge it would bear
if all the ligands were removed along with
the electron pairs that were shared with the
central atom. Neutral ligands are formally
removed in their closed-shell configurations.
Where it is not feasible or reasonable to
define an oxidation state for each individual
member of a group or cluster, it is again
recommended that the overall oxidation
level of the group be defined by a formal
ionic charge, the net charge on the
coordination entity.
Paramagnetic
Substances having a positive magnetic
susceptibility are paramagnetic. They are
attracted by a magnetic field.
Peroxidase
A heme protein (donor: hydrogen peroxide
oxidoreductase, EC class 1.1 1.1) which
catalyzes the oneelectron oxidation of a
substrate by dihydrogen peroxide.
Substrates for different peroxidases include
various organic compounds, cytochrome C,
halides, and Mn2+.
53
Photosynthesis
A metabolic process in plants and certain
bacteria, using light energy absorbed by
chlorophyll and other photosynthetic
pigments for the reduction of CO2 followed
by the formation of organic compounds.
Plastocyanin
An electron transfer protein, containing a
type 1 copper site, involved in plant and
cyanobacterial photosynthesis, which
transfers electrons to Photosystem I.
Polydentate
See chelation, donor atom symbol.
Porphyrin
A macrocyclic molecule that contains four
pyrrole rings linked between the alpha
positions of the pyrrole rings. Porphyrins
coordinated to a metal ion.
Primary
structure
The amino-acid sequence of a protein or
nucleotide sequence of DNA or RNA.
Protoporphyrin
The porphyrin ligand of heme b. Heme b is
a Fe(II) porphyrin complex readily isolated
54
from the hemoglobin of beef blood, but also
found in other proteins including other
hemoglobins, myoglobins, cytochromes P450, catalases, peroxidases as well as b type
cytochromes. Protoporphyrin IX contains
four methyl groups in positions 2, 7, 12 and
18, two vinyl groups in positions 3 and 8
and two propionic acid groups in positions
13 and 17.
Quaternary
structure
The level of structural organization in
oligomeric proteins (i.e., those composed of
more than one subunit) represented by the
number and arrangement of the subunits and
the interactions between them.
Radical
A molecular entity possessing one or more
unpaired electrons, formerly often called
“free radical”. A radical may be charged,
positively (radical cation) or negatively
(radical anion). Paramagnetic metal ions are
not normally regarded as radicals.
Redox potential
Any oxidation-reduction (redox) reaction
can be divided into two half reactions: one
in which a chemical species undergoes
oxidation and one in which another chemical
species undergoes reduction. If a
halfreaction is written as a reduction, the
driving force is the reduction potential. If the
half-reaction is written as oxidation, the
driving force is the oxidation potential
related to the reduction potential by a sign
55
change. So the redox potential is the
reductiodoxidation potential of a compound
measured under standard conditions against
a standard reference half-cell. In biological
systems the standard redox potential is
defined at pH = 7.0 versus the hydrogen
electrode and partial pressure of hydrogen =
1 bar.
Ribonucleic acids Linear polymer molecules composed of a
chain of ribose units linked between
(RNA)
positions 3 and 5 by phosphodiester groups.
The bases adenine or guanine (via atom N-9)
or uracil or cytosine (via atom N-l),
respectively, are attached to ribose at its
atom C-1 by -N-glycosidic bonds (see
nucleotides). The three most important types
of RNAs in the cell are: messenger RNA
(mRNA), transfer RNA (tRNA), ribosomal
RNA (rRNA).
Rieske ironsulfur protein
An iron-sulfur protein of the
mitochondria1 respiratory chain, in which
the (2Fe-2S] cluster is coordinated to two
sulfur ligands from cysteine and two
imidazole ligands from histidine. The term is
also applied to similar proteins isolated from
photosynthetic organisms and
microorganisms, and other proteins
containing [2Fe-2S] clusters with similar
coordination.
Rubredoxin
An iron-sulfur protein without acid-labile
56
sulfur, in which an iron center is
coordinated by four sulfur containing
ligands, usually cysteine. The function,
where known, is as an electron carrier.
Secondary
structure
Level of structural organization in proteins
described by the folding of the polypeptide
chain into structural motifs such as alpha
helices and beta sheets, which involve
hydrogen bonding of backbone atoms.
Secondary structure is also formed in
nucleic acids, especially in single-stranded
RNA’s by internal base pairing.
Sequence
The order of neighbouring amino acids in a
protein or the purine and pyrimidine bases in
RNA or DNA. See also primary structure.
Soft acid
See hard acid.
Soft base
See hard base.
Solvation
Any stabilizing interaction of a solute (or
solute moiety) and the solvent or a similar
interaction with solvent of groups of an
insoluble material (i.e., the ionic groups of
an ion-exchange resin). They generally
involve electrostatic forces and Van der
Waals forces, as well as chemically more
specific effects such as hydrogen bond
formation.
Solvolysis
Reaction with a solvent involving the
57
rupture of one or more bonds in the reacting
solute.
Stability constant An equilibrium constant that expresses the
propensity of a species to form from its
component parts. The larger the stability
constant, the more stable is the species. The
stability constant (formation constant) is the
reciprocal of the instability constant
(dissociation constant).
Stable
Stable is a term describing a system in a
state of equilibrium corresponding to a local
minimum of the appropriate thermodynamic
potential for the specified constraints on the
system. Stability cannot be defined in an
absolute sense, but if several states are in
principle accessible to the system under
given conditions, that with the lowest
potential is called the stable state, while the
other states are described as metastable.
Unstable states are not at a local minimum.
Transitions between metastable and stable
states occur at rates which depend on the
magnitude of the appropriate activation
energy barriers which separate them.
Stereochemical
Refers to the three-dimensional view of a
molecule either as such or in a projection.
Stock number
See oxidation number.
Substrates
(1) A chemical species of particular interest,
58
the reaction of which with some other
chemical reagent is under observation (e.g. a
compound that is transformed under the
influence of a catalyst).
(2) The chemical entity whose conversion to
a product or products is catalyzed by
enzymes.
(3) A solution or dry mixture containing all
ingredients which are necessary for the
growth of a microbial culture or for product
formation.
(4) A component in the nutrient medium,
supplying the organisms with carbon (Csubstrate), nitrogen (N- substrate) etc.
Superoxide
dismutases
(SOD)
Enzymes which catalyze the dismutation
reaction of superoxide anion to dihydrogen
peroxide and dioxygen. The enzymes have
active sites containing either copper and zinc
(Cd/Zn superoxide dismutase), or iron (Fesuperoxide dismutase), or manganese (Mnsuperoxide dismutase).
Tertiary
structure
The overall three-dimensional structure of a
biopolymer. For proteins this involves the
side chain
interactions and packing of secondary
structure motif. For nucleic acids this may
be the packing of stemloops or supercoiling
of double helices.
Trace elements
Elements required for physiological
functions in very small amounts that vary for
59
different organisms. Included among the
trace elements are Co, Cu, F, Fe, I, Mn, Mo,
Ni, Se, V, W, and Zn. Excess mineral intake
may produce toxic symptoms.
Transferrin
An iron-transport protein of blood plasma,
which comprises two similar iron-binding
domains with high affinity for Fe(III).
Similar proteins are found in milk
(lactoferrin) and eggs (ovotransferrin).
Transition
element
Transition element is an element whose
atom has an incomplete d-sub-shell, or
which gives rise to a cation or cations with
an incomplete d-sub-shell. The First
Transition Series of elements is Sc, Ti, V,
Cr, Mn, Fe, Co, Ni, Cu. The Second and
Third Transition Series are similarly derived
: these include the lanthanoids (lanthanides)
and actinoids (actinides) respectively which
are designated inner (or f ) transition
elements of their respective Periods in the
Periodic Table.
Type 1, 2, 3
copper
Different classes of copper-binding sites in
proteins, classified by their spectroscopic
properties as Cu(II). In type 1, or blue
copper centers the copper is coordinated to
at least two imidazole nitrogens from
histidine and one sulfur from cysteine. They
are characterized by small copper hyperfine
couplings and a strong visible absorption in
the Cu(II) state. In type 2, or non-blue
60
copper sites, the copper is mainly bound to
imidazole nitrogens from histidine. Type 3
copper centers comprise two spin-coupled
copper ions, bound to imidazole nitrogens.
Zinc finger
A domain, found originally in certain DNAbinding proteins and subsequently in other
classes of proteins, comprising a helix-loop
structure in which a zinc ion is coordinated
to 2-4 cysteine sulfurs, the remaining
ligands being histidines. In many proteins of
this type the domain is repeated several
times.
Zwitterionic
compound
A neutral compound having electrical
charges of opposite sign, delocalized or not,
on adjacent or nonadjacent atoms.
61
62
LECTURE NOTES
63
64
CHAPTER I
Introduction to Bioinorganic Chemistry
Bioinorganic chemistry is a leading discipline at the
interface of chemistry and biology. A lot of biological processes
require metal ions. Among them, there are metabolism, nitrogen
and oxygen fixation, photosynthesis, nerve transmission, and
many others. Metals are being introduced into living bodies as
diagnostic probes and drugs.
Thus, bioinorganic chemistry is a field that examines the
role of metals in biology. Bioinorganic chemistry includes the
study of both natural phenomena such as the behavior of
metalloproteins as well artificially introduced metals, including
those that are non-essential, in medicine and toxicology.
Bioinorganic chemistry is important in realizing the implications
of electron-transfer proteins, substrate bindings and activation,
atom and group transfer chemistry, as well as metal properties in
biological chemistry.
Major element content in living bodies differ from those
in the Earth’ rust but is in a good correlation with that in the sea
water:
65
Chemical elements essential to life forms can be divided
into the following:
(i) Bulk elements: C, H, N, O, P, S;
(ii) Macrominerals and ions: Na+, K+, Mg2+, Ca2+, Cl-,
PO43-, SO42-;
(iii) Trace elements: Fe2+/3+, Zn2+, Cu2+;
(iv) Ultratrace elements comprises of:
(a) non-metals: F, I, Se, Si, As, B
(b) metals: Mn, Mo, Co, Cr, V, Ni, Cd, Sn, Pb, Li.
About 99% of mammals' mass are the elements
carbon, nitrogen, calcium, sodium, chlorine, potassium,
hydrogen, phosphorus, oxygen and sulfur. The organic
compounds (proteins, lipids and carbohydrates) contain
the majority of the carbon and nitrogen and most of the
oxygen and hydrogen is present as water. The entire
66
collection of metal-containing biomolecules in a cell is
called the metallome. The biological functions of some
of the metals are presented in the Table 1.
Table 1. Biological functions of some metals.
The essentiality of metallic elements is defined by the
following:
(1) A physiological deficiency appears when the
element is removed from the diet.
(2) The deficiency is relieved by the addition of that
element to the diet.
(3) A specific biological function is associated with
the element.
Some applications of medicinal relevant elements are
listed bellow (see Table 2).
67
Table 2. Medicinal applications of some metal cations
Metal ion or salt
Application
+
Li
Treatment of bipolar disorder (maniac
depression) and hypertension
3+
Gd
Contrast agent in magnetic resonance
tomography of soft tissues
BaSO4
Contrast agent for X-ray tomography. Sun
protection
99m
Tc
(a metastable γ-emitter; t1/2 = 6 h): Radio
diagnostics (bone cancer, infarct risk, …)
2+
Pt
Chemotherapy (e.g. with cisplatin cis[Pt(NH3)2Cl2]) of cancer (ovaria, testes)
+
Au
Therapy of rheumatic arthritis
Sb3+
Treatment of inflammatory skin pimples like
acne
Bi3+
Treatment of gastritis
Transition metal ions commonly are not present in a free
form, but rather coordinated (complexed) to ligands. In particular,
this applies to metal ions in the active centres of enzymes or
otherwise integrated into peptides and proteins. Examples for the
respective ligands are listed below (N-functional: peptide moiety,
porphinogenes, histidine; O-functional: tyrosinate, serinate,
glutamate and aspartate; S-functional: cysteinate and
methionine):
68
Some additional inorganic ligands are also present:
There are three major avenues of investigation in
bioinorganic chemistry. The first involves direct study of the
structure and function of ‘metallobiomolecules’, an area which is
traditionally that of a biochemist. The second major avenue
involves an indirect approach, commonly the domain of the
inorganic or organic chemists. The third involves the addition of
metal ions or complexes as probes to biochemical structure and
function.
69
Basic concepts of the method of molecular orbitals
(МО) in description of covalent and coordinate
bonds.
According to МО method, the molecule consists of a set
of nuclei and electrons disposed on molecular orbitals.
Main tenets of the MO method:
1. Each electron in a molecule occupies a certain energetic
level (molecular orbital, MO) which is characterised by a
molecular -function and a corresponding set of quantum
numbers.
2. The total number of formed МО equals the number of
initial atomic orbitals.
3. Filling of MO occurs according to all the principles
presented for atomic orbitals.
4. МО it is considered as a linear combination of atomic
orbitals (MO – LCAO).
Let's consider, for example, formation of molecule АВ.
Valence electrons of each atom are on p-orbitals.
If the wave function of the isolated atom A is А, and for
the atom B it is В, so according to the МО method:
±АВ=С1А±С2В
where С1 and С2 are coefficients considering the income of each
atom in formation of a molecular orbital.
70
А
_
E
(ab*.)
В

А
В
+
А
В
(b)
The new molecular orbital with a lower energy (b) is
known as a bonding orbital. As its energy is lower than the
energy of an atomic orbital, electrons on it stabilize the molecule.
The MO with a higher energy (ab*) is called an
antibonding orbital. Electrons on it tend to destroy the molecule.
Stability of a molecule is described by a bond order, B.O.
B.O. = ½ (No of bonding electrons – No of antibonding
electrons)
If B.O. = 0 the number of electrons on binding orbitals
equals the number of electrons on antibinding electrons. Such
molecule is unstable and breaks up to initial atoms (does not
exist).
Conditions for formation of MO from AO are the
following:
(i) close values of energies of overlapping atomic orbitals;
(ii) considerable overlap of AO (formation of  and -types
of MO);
(iii) identical spatial disposition of AO (рх - рх, instead of
рх - рz).
On level power molecular orbitals two-nuclear molecules
settles down in a following order:
71
s(1s)s(1s)s(2s)s*(2s)x(2px)z(2pz)=y(2py)z*(2pz)
=y*(2py)x*(2px)
Examples of the description of molecules using the MO
method (energy diagrams of molecules and molecular ions):
72
73
74
75
76
Coordination compounds (definition).
Coordination compounds, or complexes, are integral
molecular or ionic units consisting of a central metal ion (or
atom), bonded to a defined number of ligands in a defined
geometrical arrangement. The ligands can be ions or (induced)
dipolar molecules. Each ligand provides a free electron pair, i.e.
the ligands are Lewis bases, while the metal in the coordination
centre is the Lewis acid. The bonding can thus be described in
terms of Lewis acid/Lewis base interaction. Other descriptions of
the bonding situation are: (i) donor bond; (ii) coordinative
covalent bond, often denoted by L→M, where L = ligand and M
= metal.
Complexes tend to be stable when the overall electron
configuration at the metal centre (the sum of metal valence
electrons plus electron pairs provided by the ligands) is 18 (or 16
for the late transition metals).
Structure of complex compounds
In a molecule of a complex compound, one of the atoms,
generally positively charged, occupies the central site (central
ion or complexing agent). Oppositely charged ions or neutral
molecules called ligands are coordinated around the central ion.
The complexing agent and ligands form inner sphere of a
complex compound. It is characterized by coordinate bonds
which are formed while overlapping of empty p- and d-orbitals of
a central ion and orbitals containing lone electron pairs of
ligands. The ions in the outer sphere are mainly bonded to the
complex ions by forces of electrostatic interaction (ionic bonds).
The total number of coordinate bonds formed by the
complexing agent is known as coordination number of the
central ion. It mainly depends upon the charge of the complexing
agent (for monocharged ions it usually equals 1, for discharged
77
ions - 4 or 6, for tricharged - 6 and above), and the size of an ion
(the larger the central ion, the greater its coordination number is,
for lanthanides and actinides it can reach to 12)..
Ligands possess the property of dentacity. In accordance
with the number of coordinate bonds formed by a ligand with the
central ion, the ligand may be a monodentate, bidentate, or
polydentate. Dentacity is defined by number lone electronic pairs
in a molecule of a ligand and their mutual spatial disposition. For
example, the ammonia molecule NH3 has one lone electronic pair
belonging to the N-atom therefore ammonia is a monodentate
ligand. The water molecule has two lone electronic pairs, and
chloride-ion has them four. However because valence orbitals of
oxygen and chlorine are in the sp3-type of hybridization and are
located under a corner 10928 ’ they cannot form chemical bonds
with the same central ion, therefore such лиганды are
monodentate except the case of polynuclear complexes where
they act as bidentate bridging ligands.
Example: Na2 [Cu2Cl6]
Cl
Na2
Cl
Cl
Cu
Cu
Cl
Cl
Cl
Dicharged acidic anions such as SO32–, C2O42– ets. are
usually bidentate chelating ligands.
Example: Na2[Cd(SO3)2]
78
O
Na2
O S
O
Cd
O
S O
O
Properties

Multidentate ligands are much stronger complexes
than monodentate ligands;
 Chelates remain stable even at very dilute
concentrations (less dissociation);
 Chelate effect: increase of entropy
G = H – TS, H for multi- and monodentate
complexes
Cu(H2O)42+ + 4NH3  [Cu(NH3)4]2+ + 4H2O
Cu(H2O)42+ + N4X  [Cu(N4X)]2+ + 4H2O
 Chelate therapy (detoxification).
79
Electronic structure of complex ions.
I. Valence bond theory (Pauling).
Interaction of lone electronic pairs of ligands with empty
valence orbitals of the central ion of different types leads to their
hybridization. For example, the electronic structure of a complex
ion [Cu(NH3)4]2+ can be reflected as following:
2 2 6 2
6 10 1
29Cu: 1s 2s 2p 3s 3p 3d 4s
2+
2 2 6 2
6 9 0 0 0
29Cu : 1s 2s 2p 3s 3p 3d 4s 4p 4d





3d
4s
4p
4d
4 :NH3
Interaction of one s- and three p-orbitals leads to the sp3hybridization of the central ion (tetrahedral complex).
Entering of lone electronic pairs of ligands into valence
orbitals of the central ion leads to their interaction with the
electrons of 3d-orbitals. This interaction is defined by degree of
penetration of electrons of ligands on empty orbitals of metallic
cations. In connection of force of interaction, ligands may be
arranged in a spectrochemical series and are devided into ligands
of weak and strong field:
CO, CN– > NO2– > NH3
Ligands of a strong field
> SCN–
> H2O
> OH–- > F–
Ligands of a weak field
> Cl–
Lone electronic pairs of ligands of a strong field deeply
enter the valence electronic shell of the central ion and cause
pairing of electrons of the 3d-subshell.
80
As a result, intra-orbital low-spin complexes are formed
(coordination bonds are formed with participation of internal 3dorbitals, the formed complex has no or few unpaired electrons):
2 2 6 2
6 7 2
27Co: 1s 2s 2p 3s 3p 3d 4s
3+
2 2 6 2 6 6 0 0 0
27Co : 1s 2s 2p 3s 3p 3d 4s 4p 4d





3d
4s
4p
4d
4s
4p
4d
[Co(NH3)6]3+



3d
6 :NH3
d2sp3-hybridization, octahedral complex.
Lone electronic pairs of ligands of a weak field slightly
interact with 3d-electrons of the central ion and do not cause their
pairing. As a result, outer-orbital high-spin complexes are
formed:
81
[CoF6]3–




3d

4s
4p
4d
6 :F-
sp3d2-hybridization, octahedral complex.
Crystal Field Theory (Electrostatic guide to splitting of
d-levels).

Ligand field splitting 0, LFSE (Ligand Field
Splitting Energy);
 Spectrichemical series (0 increases):
I- < Br- < S2- < Cl- < NO3- < OH- < H2O < NH3 < en <
NO2- < CN- < CO;
 Metal 0 increases with increasing oxidation
number and down a group:
Mn(II) < Ni(II) < Fe(II) < V(II) < Fe(III) < Co(III) < Mn(IV) <
Mo(III) < Pd(IV) < Pt(IV);
 Electronic configuration of a central atom depends
on LFSE and P (pairing energy);
 Jahn Teller distortion: remove of degeneracy,
increase of LFSE:
i)
splitting eg (dz2 lower energy) and t2g (dxy
higher energy;
ii)
square coordination: dxy higher energy than
dz2.
82
As an example, the versatility of iron as a function of its
environment is is discussed. The ligand field can strongly alter
the structural and ligand exchange properties of the metal ion
(Figure 1). The ligand field can also alter the redox properties.
For high-spin ferric ion, as found in the aquo complex or in many
other complexes (including the class of microbial iron-transport
agents called siderophores, to be discussed later), the
coordination geometry is octahedral or pseudo-octahedral.
Figure 1. Versatility of Fe coordination compounds.
83
In the relatively weak ligand field (high-spin ground
state), the complex is highly labile. In a strong ligand field, such
as an axially ligated porphyrin complex of ferric ion, or the
simple example of the ferrocyanide anion, the low-spin complex
is exchange-inert. Similarly, the high-spin octahedral ferrous
complexes are exchange-labile, but the corresponding axially
ligated porphyrin complexes, or the ferrocyanide complexes, are
spin-paired (diamagnetic) and ligand exchange-inert. Large,
bulky ligands or constrained ligands, such as those provided by
metalloprotein and enzyme sites, can cause a tetrahedral
environment, in which both ferrous ion and ferric ion form highspin complexes.
Example: FeII (d6): In an octahedral (Oh) field, the
degeneracy of the five d-orbitals is lifted. Depending on the
strength of the ligand field, the ligand field stabilisation energy
(i.e. the energy set free as all of the electrons are accommodated
in the orbitals of lower energy) can be (i) less and (ii) more than
the energy needed for electron pairing. In the case of (i), i.e. aqua
ligands, a high-spin complex is formed; in the case of (ii), i.e.
cyanido ligands, a low-spin complex is formed. Asymmetrically
occupied orbital sets, as in the case of [Fe(H2O)6]2+, result in
further stabilisation through symmetry lowering: Jahn Teller
distortion:
84
85
Pearson’s concept of hard and soft interactions.









Lewis concept: metal ions are acids because they accept
electrons. Ligands are bases because they donate
electrons.
Hard acids tend to form complexes with hard bases (ionic
bonds)
Soft acids tend to form complexes with soft bases
(covalent bonds)
Hard acids: H+, Li+(> Na+…), Cr6+(> Cr3+)
Intermediate acids: Fe2+, Mn2+, Cu2+, Zn2+
Soft acids: Au+(> Ag+, Cu+), Hg2+, Pt2+
Hard bases: F-, OH-, NH3, PO43-(> HPO42-), MoO42Intermediate bases: ClSoft bases: I-(> Br-…), S2-(> HS-, > H2S), AsS2-
Chelate effect
(Stabilisation of complexes by multidentate ligands).
The chelate effect is an entropic effect (high entropy =
high disorder [increase of particle number]).
Example: The complex formed between the siderophore
enterobactin:
86
(ent6-, a hexadentate ligand) and Fe3+ is particularly stable:
[Fe(H2O)6]3+ + ent6- → [Fe(ent)]3- + 6H2O.
87
CHAPTER II.
Transition Metal Chemistry
Living organisms store and transport transition metals
both to provide appropriate concentrations of them for use in
metalloproteins or cofactors and to protect themselves against the
toxic effects of metal excesses; metalloproteins and metal
cofactors are found in plants, animals, and microorganisms. The
form of the metals in living bodies is always ionic, but the
oxidation state can vary, depending on biological needs.
Transition metals for which biological storage and transport are
significant are, in order of decreasing abundance in living
organisms: iron, zinc, copper, molybdenum, cobalt, chromium,
vanadium, and nickel.
Iron
Iron takes over a central role in biological events. On the
one hand, this is due to its general availability (iron is abundant
and ubiquitous in the geo- and biospheres), on the other hand,
iron has specific and “biologically suitable” properties otherwise
not (or less) available with other transition metals:
(1) Ease of change between the oxidation states +II and
+III (and disposability also of the oxidation states +IV and +V);
(2) Formation of hexaaqua cations in water; these
hexaaqua cations are Brønsted acids;
(3) Tendency to form oligo- and polymers by
condensation;
(4) Easy change between high- and low-spin states in
ligand fields of medium strength (spin cross-over);
88
(5) Flexibility with respect to the nature of the donating
ligand function, the coordination number and coordination
geometry.
In aqueous solutions, the redox potential for the pair
Fe /Fe at pH = 7 demonstrates that FeII is easily oxidised to
FeIII under aerobic conditions:
Fe2+
Fe3+ + e-; E = -0.23 V (at pH 7)
2+
3+
Hexaaquairon(III) ions are cationic Brønstedt acids:
[Fe(H2O)6]3+ + H2O
[Fe(H2O)5OH]2+ + H3O+
pKS1 = 2.2
[Fe(H2O)5(OH)]2+ + H2O
[Fe(H2O)4(OH)2]+ + H3O+
pKS2 = 3.5
[Fe(H2O)4(OH)2]+ + H2O
[Fe(H2O)3(OH)3]
(= Fe(OH)3·aq + H3O+)
pKS3 = 6.0
The formation of ferric hydroxide Fe(OH)3·aq hence
already starts in weakly acidic media. The protolytic reactions are
accompanied by condensation reactions, leading to hydroxidoand oxido-bridged aggregates and finally to colloids and hardly
soluble ferric oxide hydrates:
89
While coordination, Fe-ions posses the affinity to Odonating groups of the catecholate and hydroximate-like ligands:
90
The formation of coordinate bonds with N- and Sdonating atoms occurs in hemoglobine (a), cytochrom c (b) and
rubredoxin (c) structures:
91
(a)
(b)
92
(c)
The distribution of specific iron complexes in living
organisms depends strongly on function. For example, although
there are many different iron complexes in the average human,
the relative amounts of each type differ more than 650-fold
(Table 3). The total amount of iron in humans is quite large,
averaging more than three and up to five grams for a healthy
adult. Most of the iron is present as hemoglobin, the plasma
oxygen-transport protein. A much smaller amount of iron is
present in myoglobin, a muscle oxygen-storage protein.
Other examples of iron-containing proteins and their
functions are included in Table 3.
93
Table 3. Average human Fe distribution
Different iron coordination environments significantly
changes the redox-potential of the Fe3+/Fe2+ redox-pair (Table 4).
Table 4. Fe redox potentials
94
Chemical properties of zinc, copper, vanadium,
chromium, molybdenum, and cobalt
For zinc there is only the +2 oxidation state, and the
hydrolysis of this ion is not a limiting feature of its solubility or
transport. Zinc is an essential element for both animals and
plants.The Zn2+ cation is an efficient Lewis acid, making it a
useful catalytic agent in hydroxylation and other enzymatic
reactions. The metal also has a flexible coordination geometry,
which allows proteins using it to rapidly shift conformations to
perform biological reactions. Two examples of zinc-containing
enzymes are carbonic anhydrase and carboxypeptidase, which are
vital to the processes of carbon dioxide (CO2) regulation and
digestion of proteins, respectively. In vertebrate blood, carbonic
anhydrase converts CO2 into bicarbonate and the same enzyme
transforms the bicarbonate back into CO2 for exhalation through
the lungs. Without this enzyme, this conversion would occur
about one million times slower at the normal blood pH of 7 or
would require a pH of 10 or more. The non-related β-carbonic
anhydrase is required in plants for leaf formation, the synthesis of
indole acetic acid (auxin) and alcoholic fermentation.
Zinc serves a purely structural role in zinc fingers, twists
and clusters (Figure 2).
95
Figure 2. Zinc fingers help read DNA sequences.
Zinc fingers form parts of some transcription factors,
which are proteins that recognize DNA base sequences during the
replication and transcription of DNA. Each of the nine or ten Zn2+
ions in a zinc finger helps maintain the finger's structure by
coordinately binding to four amino acids in the transcription
factor. The transcription factor wraps around the DNA helix and
uses its fingers to accurately bind to the DNA sequence.
Copper, like all other transitional metals, forms
coordination complexes. In aqueous solution, copper(II) exists as
[Cu(H2O)6]2+. This complex exhibits the fastest water exchange
rate (speed of water ligands attaching and detaching) for any
transition metal aquo complex. Adding aqueous sodium
hydroxide causes the precipitation of light blue solid copper(II)
hydroxide. A simplified equation is: Cu2+ + 2 OH− → Cu(OH)2
96
Aqueous ammonia results in the same precipitate. Upon
adding excess ammonia, the precipitate dissolves, forming
tetraamminecopper(II):
Cu(H2O)4(OH)2 + 4 NH3 → [Cu(H2O)2(NH3)4]2+ +
2 H2O + 2 OH−
Many other oxyanions form complexes; these include
copper(II) acetate, copper(II) nitrate, and copper(II) carbonate.
Copper(II) sulfate forms a blue crystalline pentahydrate, which is
the most familiar copper compound in the laboratory. It is used in
a fungicide called the Bordeaux mixture.
Polyols, compounds containing more than one alcohol
functional group, generally interact with cupric salts. For
example, copper salts are used to test for reducing sugars.
Specifically, using Benedict's reagent and Fehling's solution the
presence of the sugar is signaled by a color change from blue
Cu(II) to reddish copper(I) oxide. Schweizer's reagent and related
complexes with ethylenediamine and other amines dissolve
cellulose.[34] Amino acids form very stable chelate complexes
with copper(II). Many wet-chemical tests for copper ions exist,
one involving potassium ferrocyanide, which gives a brown
precipitate with copper(II) salts.
Copper proteins are proteins that contain one or more
copper ions as prosthetic groups. The metal centers in the copper
proteins can be classified into several types:
Type I copper centers (T1Cu) (Figure 3) are
characterized by a single copper atom coordinated by two
97
histidine residues and a cysteine residue in a trigonal planar
structure, and a variable axial ligand.
Figure 3. Structure of poplar plastocyanin
In class I T1Cu proteins (e.g. amicyanin, plastocyanin and
pseudoazurin) the axial ligand is the sulfur of methionine,
whereas aminoacids other than methionine (e.g. glutamine) give
rise to class II T1Cu copper proteins. Azurins contain the third
type of T1Cu centres: besides a methionine in one axial position,
they contain a second axial ligand (a carbonyl group of a glycine
residue). T1Cu-containing proteins are usually called
"cupredoxins", and show similar three-dimensional structures,
relatively high reduction potentials (> 250 mV), and strong
absorption near 600 nm (due to S→Cu charge transfer), which
usually gives rise to a blue colour. Cupredoxins are therefore
often called "blue copper proteins". This may be misleading,
since some T1Cu centres also absorb around 460 nm and are
therefore green. When studied by EPR spectroscopy, T1Cu
centres show small hyperfine splittings in the parallel region of
98
the spectrum (compared to common copper coordination
compounds).
Type II copper centres (T2Cu) exhibit a square planar
coordination by N or N/O ligands. They exhibit an axial EPR
spectrum with copper hyperfine splitting in the parallel region
similar to that observed in regular copper coordination
compounds. Since no sulfur ligation is present, the optical spectra
of these centres lack distinctive features. T2Cu centres occur in
enzymes, where they assist in oxidations or oxygenations.[2]
Type III copper centres (T3Cu) consist of a pair of
copper centres, each coordinated by three histidine residues.
These proteins exhibit no EPR signal due to strong
antiferromagnetic coupling (i.e. spin pairing) between the two S =
1/2 metal ions due to their covalent overlap with a bridging
ligand. These centres are present in some oxidases and oxygentransporting proteins (e.g. hemocyanin and tyrosinase).
Binuclear Copper A centres (CuA) (Figure 4) are found
in cytochrome c oxidase and nitrous-oxide reductase (EC
1.7.99.6). The two copper atoms are coordinated by two
histidines, one methionine, a protein backbone carbonyl oxygen,
and two bridging cysteine residues.
99
Figure 4. Crystal structure of hexameric haemocyanin
Copper B centres (CuB) are found in cytochrome c
oxidase. The copper atom is coordinated by three histidines in
trigonal pyramidal geometry.
Tetranuclear Copper Z centre (CuZ) is found in nitrousoxide reductase. The four copper atoms are coordinated by seven
histidine residues and bridged by a sulfur atom.
Copper proteins have diverse roles in biological electron
transport and oxygen transportation, processes that exploit the
easy interconversion of Cu(I) and Cu(II). The biological role for
copper commenced with the appearance of oxygen in earth's
atmosphere. The protein hemocyanin is the oxygen carrier in
most mollusks and some arthropods such as the horseshoe crab
(Limulus polyphemus). Because hemocyanin is blue, these
100
organisms have blue blood, not the red blood found in organisms
that rely on hemoglobin for this purpose. Structurally related to
hemocyanin are the laccases and tyrosinases. Instead of
reversibly binding oxygen, these proteins hydroxylate substrates,
illustrated by their role in the formation of lacquers.
Copper is also a component of other proteins associated
with the processing of oxygen. In cytochrome c oxidase, which is
required for aerobic respiration, copper and iron cooperate in the
reduction of oxygen. Copper is also found in many superoxide
dismutases, proteins that catalyze the decomposition of
superoxides, by converting it (by disproportionation) to oxygen
and hydrogen peroxide: 2 HO2 → H2O2 + O2.
Several copper proteins, such as the "blue copper
proteins", do not interact directly with substrates, hence they are
not enzymes. These proteins relay electrons by the process called
electron transfer
Vanadium is often taken up as vanadate, in a pathway
parallel to phosphate. However, its oxidation state within
organisms seems to be highly variable.Unusually high
concentrations of vanadium occur in certain ascidians (the
specific transport behavior of which will be dealt with later). The
workers who first characterized the vanadium-containing
compound of the tunicate, Ascidia nigra, coined the name
tunichrome. The characterization of the compound as a
dicatecholate has been reported.
Quite a different chemical environment is found in the
vanadium-containing material isolated from the mushroom
Amanita muscaria. Bayer and Kneifel, who named and first
described amavadine, also suggested the structure shown in
Figure 5. Recently the preparation, proof of ligand structure, and
101
(by implication) proof of the complex structure shown in Figure 5
have been established.
Figure 5. A structure proposed for amavadine.
The role of chromium in biology remains even more
mysterious. In human beings the isolation of "glucose tolerance
factor" and the discovery that it contains chromium goes back
some time. This has been well reviewed by Mertz, who has
played a major role in discovering what is known about this
elusive and apparently quite labile compound. It is well
established that chromium is taken up as chromic ion,
predominantly via foodstuffs, such as unrefined sugar, which
presumably contain complexes of chromium, perhaps involving
sugar hydroxyl groups.
Studies using radioactively labeled chromium have shown
that, although inorganic salts of chromium are relatively
unavailable to mammals, brewer's yeast can convert the
chromium into a usable form.
Although chromium is essential in milligram amounts for
human beings as the trivalent ion, as chromate it is quite toxic
and a recognized carcinogen. The uptake-reduction model for
102
chromate carcinogenicity as suggested
Wetterhahn is shown in Figure 6.
by
Connett
and
Figure 6. The uptake-reduction model for chromate
carcinogenicity. Possible sites for reduction of chromate include
the cytoplasm, endoplasmic reticulum, mitochondria, and the
nucleus
Chromate is mutagenic in bacterial and mammalian cell
systems, and it has been hypothesized that the difference between
chromium in the +6 and +3 oxidation states is explained by the'
'uptake-reduction" model. Chromium(III), like the ferric ion
discussed above, is readily hydrolyzed at neutral pH and
103
extremely insoluble. Unlike Fe3+ , it undergoes extremely slow
ligand exchange. For both reasons, transport of chromium(III)
into cells can be expected to be extremely slow unless it is
present as specific complexes; for example, chromium(III)
transport into bacterial cells has been reported to be rapid when
iron is replaced by chromium in the siderophore iron-uptake
mediators. However, chromate readily crosses cell membranes
and enters cells, much as sulfate does. Because of its high
oxidizing power, chromate can undergo reduction inside
organelles to give chromium(III), which binds to small
molecules, protein, and DNA, damaging these cellular
components.
In marked contrast to its congener, molybdenum is very
different from chromium in both its role in biology and its
transport behavior, again because of fundamental differences in
oxidation and coordination chemistry properties. In contrast to
chromium, the higher oxidation states of molybdenum dominate
its chemistry, and molybdate is a relatively poor oxidant.
Molybdenum is an essential element in many enzymes, including
xanthine oxidase, aldehyde reductase, and nitrate reductase. The
range of oxidation states and coordination geometries of
molybdenum makes its bioinorganic chemistry particularly
interesting and challenging.
Common oxidation states of cobalt include +2 and +3,
although compounds with oxidation states ranging from −3 to +4
are also known. A common oxidation state for simple compounds
is +2. Cobalt(II) salts form the red-pink [Co(H2O)6]2+ complex in
aqueous solution. Addition of chloride gives the intensely blue
[CoCl4]2−.[
The principles of electronegativity and hardness–softness
of a series of ligands can be used to explain the usual oxidation
state of the cobalt. For example Co+3 complexes tend to have
ammine ligands. As phosphorus is softer than nitrogen, phosphine
104
ligands tend to feature the softer Co2+ and Co+, an example being
tris(triphenylphosphine)cobalt(I) chloride ((P(C6H5)3)3CoCl). The
more electronegative (and harder) oxide and fluoride can stabilize
Co4+ and Co5+ derivatives, e.g., caesium hexafluorocobaltate
(Cs2CoF6) and potassium percobaltate (K3CoO4).[
Figure 7. Structure of Co complexes with biomolecules
105
In the living bodies, cobalt is found in coenzyme B12
(Figure 7), which is essential for the transfer of alkyl groups from
one molecule to another in biological systems, as well as the
reduction of the ribose ring in ribonucleotides that make up RNA
to the deoxyribose ring in deoxyribonucleotides that make up
DNA.
The cobalamin-based proteins use corrin to hold the
cobalt. Coenzyme B12 features a reactive C-Co bond, which
participates in its reactions. In humans, B12 exists with two types
of alkyl ligand: methyl and adenosyl. MeB12 promotes methyl (CH3) group transfers. The adenosyl version of B12 catalyzes
rearrangements in which a hydrogen atom is directly transferred
between two adjacent atoms with concomitant exchange of the
second substituent, X, which may be a carbon atom with
substituents, an oxygen atom of an alcohol, or an amine.
Methylmalonyl coenzyme A mutase (MUT) converts MMl-CoA
to Su-CoA, an important step in the extraction of energy from
proteins and fats.
Although far less common than other metalloproteins (e.g.
those of zinc and iron), cobaltoproteins are known aside from B12.
These proteins include methionine aminopeptidase 2 an enzyme
that occurs in humans and other mammals which does not use the
corrin ring of B12, but binds cobalt directly. Another non-corrin
cobalt enzyme is nitrile hydratase, an enzyme in bacteria that are
able to metabolize nitriles.
106
CHAPTER III
Structure, properties, and functions of
metal complexes of proteins and nucleic acids
Part 1. Metalloprotein structures
Proteins are large biological molecules consisting of one
or more chains of -amino acids:
The structure of proteins includes:
•primary structure: sequence of amino acids;
•secondary structure: shapes formed within regions of
the protein;
•tertiary structure: shape of entire protein;
•quaternary structure: structures formed by interaction
of several subunit.
Proteins perform a vast array of functions within living
organisms, including catalyzing metabolic reactions, replicating
DNA, responding to stimuli, and transporting molecules from one
location to another. Proteins differ from one another primarily in
their sequence of amino acids, which is dictated by the nucleotide
sequence of their genes, and which usually results in folding of
the protein into a specific three-dimensional structure that
determines its activity.
107
It is estimated that approximately half of all proteins
contain a metal. In another estimate, about one quarter to one
third of all proteins are proposed to require metals to carry out
their functions.
Amino acids are important low molecular weight ligands
in biological systems as metals are bound to proteins through
amino acid side chains:
108
Coordination through carboxylate has three basic modes:
Characteristic affinities of some amino acids to defined
oxidation states of metals, typical coordination numbers and
coordination geometries are presented below:
Metal centers are undersaturated (bonding of substrate),
and their coordination geometries are frequently distorted. So, the
possibility of extra coordination of some exogenous ligands
109
which are not integrated into the active site of the protein exists.
The most common of them are the following: (i) water-derived
(H2O, OH-); (ii) oxygen-derived (O2, O2-, O22-, HO2-); (iii) neutral
species (CO, NO); (iv) halides and other charged species (Cl-,
PO43-).
Systematic of copper proteins
Type I (Blue Cu-proteins): trigonal coordination
geometry; ligands: 2 Cys(1-), 1 His, 1 weakly bonded Met.
Function: e- transfer; Example: Plastocyanin in the e- transfer
chain PSII→PSI.
Type II: Tetragonal coordination geometry; ligands: His,
Tyr(1-), H2O, no Cys. Function: Redox reactions; Examples:
Galactoseoxidase (RCH2OH → RCHO + 2H+ e-), Cu/Zn
superoxidedismutase (2O2- + 2H+ → O2 + H2O2).
Type III: Contains 2 cooperating Cu centers; trigonal
coordination geometry; ligands: 3 His per Cu. Intensely blue in
the oxidized form (O22- →Cu2+ LMCT). Function: Transport and
transfer (to a substrate) of oxygen; examples: haemocyanin
(Figure 8); tyrosinase (Tyr + ½O2 + 2e- → DOPA).
Figure 8. Reversible uptake and release of oxygen by
haemocyanins
110
Ceruloplasmin, important for the absorption of iron, is a
Cu protein containing 7 Cu centers representing types I, II and
III. Nitritereductase contains type II (substrate activation) and
type I Cu centres (e- transfer). Among the others, there are CuA
und CuB in cytochrom-c oxidase (Figure 9).
Figure 9. Organisation of the redox-active centres of cytochromec oxidase (left). Oxygen activation and reduction occurs at the
dinuclear CuB⋅⋅⋅Cyt-a3 pair
(see expansion to the right).
111
Iron-Sulfur Proteins.
Transition-metal/sulfide sites, especially those containing
iron, are present in all forms of life and are found at the active
centers of a wide variety of redox and catalytic proteins. These
proteins include simple soluble electron-transfer agents (the
ferredoxins), membrane-bound components of electron-transfer
chains, and some of the most complex metalloenzymes, such as
nitrogenase, hydrogenase, and xanthine oxidase.
The most important (in the sense that they are more
generally used) members of this family are collated in Figure 10.
(1) Rubredoxins contain one iron centre tetrahedrally
coordinated to four cysteinates.
(2) [2Fe,2S] ferredoxines, with two iron centers,
constitute twoedge-bridged FeS4 tetrahedra. The bridging sulphur
functions are inorganic sulphide S2-, the remaining ligands are
cysteinates.
(3) [4Fe,4S] ferredoxins have a cubane structure. The four
trebly bridging functions are again sulfide, also termed labile
sulfur because they can be converted to volatile H2S with acids.
The mean oxidation state in the reduced form is 2.25, in the
oxidized form 2.5, the redox potential is typically around -200
mV.
(4) HiPIPs (High Potential Iron Proteins) are identical to
the [4Fe,4S] ferredoxins in as far as the core structure is
concerned. However, the mean oxidation state in the reduced
form is 2.5, in the oxidized from 2.75, and the redox potential is
typically around +300 mV. Along with these “classical”
iron/sulfur clusters, others are know, in which one iron centre is
missing ([3Fe,4S] ferredoxins), or where two [4Fe,4S]
ferredoxins form double-cubanes, or where a fifth ligand (Ser or
His) is coordinated to one of the iron centers.
112
Figure 10. The iron centres of the classical (and more frequently
used) iron-sulphur proteins. SR = cysteinate(1-).
Iron/sulfur proteins are for example included in the
mitochondrial respiratory chain. The reduction of O2 to H2O takes
place step by step in order to prevent damage to cellular
constituents by the burst of energy liberated in a single step. The
reaction cascade is termed respiratory chain, which takes place in
the mitochondria, and serves the generation of energy. The
process is illustrated in the Figure 11.
113
Figure 11. Reaction cascade in the mitochondrial respiratory
chain (shortened).
Iron sulfide proteins are also involved involved in electron
transfer processes. The Figure 12 shows the ranges of redox
behavior known for Fe-S centers. Clearly, the Fe-S systems can
carry out low-potential processes. The rubredoxins cover the midpotential range, and the HiPIPs are active in the high-potential
region.
114
Figure 12. A schematic diagram of the redox potential of the
various FeS centers in comparison with other known redox
centers.
Part 2. Nucleic Acids.
Nucleic acids are large biological molecules essential for
all known forms of life. They include DNA (deoxyribonucleic
acid) and RNA (ribonucleic acid). Together with proteins, nucleic
acids are the most important biological macromolecules; each is
found in abundance in all living things, where they function in
encoding, transmitting and expressing genetic information.
Nucleic acids are linear polymers (chains) of nucleotides.
Each nucleotide consists of three components: a purine or
pyrimidine nucleobase (sometimes termed nitrogenous base or
115
simply base), a pentose sugar, and a phosphate group. The
substructure consisting of a nucleobase plus sugar is termed a
nucleoside. Nucleic acid types differ in the structure of the sugar
in their nucleotides - DNA contains 2'-deoxyribose while RNA
contains ribose (where the only difference is the presence of a
hydroxyl group). Also, the nucleobases found in the two nucleic
acid types are different: adenine, cytosine, and guanine are found
in both RNA and DNA, while thymine occurs in DNA and uracil
occurs in RNA.
The sugars and phosphates in nucleic acids are connected
to each other in an alternating chain (sugar-phosphate backbone)
through
phosphodiester
linkages.[10]
In
conventional
nomenclature, the carbons to which the phosphate groups attach
are the 3'-end and the 5'-end carbons of the sugar. This gives
nucleic acids directionality, and the ends of nucleic acid
molecules are referred to as 5'-end and 3'-end. The nucleobases
are joined to the sugars via an N-glycosidic linkage involving a
nucleobase ring nitrogen (N-1 for pyrimidines and N-9 for
purines) and the 1' carbon of the pentose sugar ring.
The structures of the components of two principal nucleic
acids RNA and DNA are presented on the Figure 13.
116
Figure 13. Nucleic acids components and structure.
117
Coordination of Nucleic Acids with Metals.
Most prevalent among covalent complexes with DNA are
those involving coordination between soft metal ions and
nucleophilic positions on the bases. For example, the structure of
the fragment of cis-(NH3)2Pt-dGpG is presented on the Figure 14.
It is evident that the platinum center coordinates to the N7
position of the guanine bases. In terms of interactions with the
full polynucleotide, it is likely that the cis-diammineplatinum
center, with two coordination sites available, would yield an
intrastrand crosslink between neighboring guanine residues on a
strand.
Figure 14. Possible coordination modes of nucleic acids
Base binding at the purine N7 position is, of course, not
limited to soft metal ions such as Pt(II), Pd(II), and Ru(II).
118
Coordination at these sites has been evident also with first-row
transition-metal ions such as Cu(II) and Zn(II).
Transition-metal ions with decreasing softness are capable
of coordinating also to the phosphate oxygen atoms. The ionic
versus covalent character of these complexes clearly depends on
the metal ions involved. The preference for phosphate over base
association was found to decrease in the order Mg(II) > Co(II)
>Ni(II) >Mn(II) >Zn(II) >Cd(II) >Cu(II). This series arises from
examination of DNA helix-melting temperatures, since base
interactions in general should destabilize the helical form [except
where interstrand crosslinking occurs, as may happen with
Ag(I)], whereas phosphate coordination and neutralization would
increase the helix stability and hence the melting temperature.
Also of interest, but less common, are covalent
interactions with the sugar moiety. Although the pentose ring in
general provides a poor ligand for metal ions, osmate esters can
form quite easily across the C2'-C3' positions in the ribose rings.
This particular interaction has been suggested as a basis for heavy
metal staining of RNA. In fact, OsO4 is not restricted in its
reactivity with the sugar positions.
Intercalation and hydrogen bonding.
Important interactions of metal complexes with
polynucleotides are not restricted to those involving direct
coordination of the metal center to the polymer. Instead, an
abundance of highly selective interactions arise from an ensemble
of weaker noncovalent interactions between the ligands of
coordinatively saturated metal complexes and the nucleic acid.
Two primary examples of noncovalent association are given by
metallointercalation and hydrogen-bonding interactions of
coordinated ligands (Figure 15). Planar aromatic heterocyclic
ligands such as phenanthroline and terpyridine can stack in
between the DNA base pairs, stabilized through dipole-dipole
119
interactions. Here, depending on the complex and its extent of
overlap with the base pairs, the free energy of stabilization can
vary from ~2 to 10 kcal. Nonintercalative hydrophobic
interactions of coordinated ligands in the DNA grooves also can
occur, as we will see. Hydrogenbonding interactions of
coordinated ligands with the polynucleotide are quite common,
and arise in particular with the phosphate oxygen atoms on the
backbone. With cobalt hexaammine, for example, hydrogen
bonding to an oligonucleotide occurs between the ammine
hydrogens and both phosphate oxygen atoms and purine bases.
A mix of covalent and noncovalent interactions is also
possible. With cisdiammineplatinum(II) coordinated to the
guanine N7 position, the ammine ligands are well-poised for
hydrogen-bonding interactions with the phosphate backbone. The
steric constraints on the molecule must be, however, considered.
120
(a)
(b)
Figure 15. Examples of intercalation (a) and hydrogen
bonding (b) in the processes of interaction of metal ions with
nucleic acids
121
Fundamental Reactions with Nucleic Acids
Complexes.
The reactions of transition-metal complexes with
polynucleotides generally fall into two categories:
(i) those involving a redox reaction of the metal complex
that mediates oxidation of the nucleic acid; and
(ii) those involving coordination of the metal center to the
sugar-phosphate backbone so as to mediate hydrolysis of the
polymer.
1. Redox chemistry.
The simplest redox reaction with polynucleotides one
might consider as an illustration is the Fenton reaction, which
indirectly promotes DNA strand scission through radical
reactions on the sugar ring. The reaction with Fe(EDTA)2 is
shown in Figure 16.
As do other redox-active divalent metal ions, ferrous ion,
in the presence of hydrogen peroxide, generates hydroxyl
radicals, and in the presence of a reductant such as
mercaptoethanol, the hydroxyl radical production can be made
catalytic. Although ferrous ion itself does not appear to interact
appreciably with a nucleic acid, especially when chelated in an
anionic EDTA complex and repelled by the nucleic-acid
polyanion, the hydroxyl radicals, produced in appreciable
quantities catalytically, attack different sites on the sugar ring,
indirectly yielding scission of the sugar-phosphate backbone.
122
Figure 16. Scheme of a redox-reaction with Fe(EDTA)2
Metal ions can also be used to generate other oxidizing
intermediates in aerated aqueous solution, such as superoxide ion
and singlet oxygen.
2. Hydrolytic chemistry.
Hydrolysis reactions of nucleic acids mediated by metal
ions are important elements in natural enzymatic reactions;
123
chemists would like to exploit them in the design of artificial
restriction endonucleases. Hydrolysis reactions of the
phosphodiester linkage of polynucleotides appear preferable to
redox-mediated cleavage reactions, since in the hydrolytic
reaction all information is preserved.
In redox cleavage by sugar oxidation, for example, both a
sugar fragment and free nucleic-acid base are released from the
polymer, and, in contrast to hydrolytic chemistry, the direct
relegation of the fragments becomes practically impossible.
Metal ions can be effective in promoting hydrolysis of the
phosphodiester, since they can function as Lewis acids, polarizing
the phosphorus-oxygen bond to facilitate bond breakage, and can
also deliver the coordinated nucleophile to form the
pentacoordinate phosphate intermediate.
Last, it must be mentioned that metal coordination to the
purine N7 position can also indirectly promote strand cleavage,
although not through direct hydrolytic reaction on the sugarphosphate backbone. Metal ions such as Pd2+ and Cu2+, through
coordination at N7, promote depurination. The depurinated site
then becomes easily susceptible to hydrolysis upon treatment
with mild base:
124
Hydrolysis reactions catalyzed by metal ions and
complexes:
(a) Illustration of a phosphate ester hydrolysis in a
binuclear model complex catalyzed by coordinated cobaltic ions,
with one metal ion functioning as a Lewis acid and the other
functioning to deliver the coordinated hydroxide.
(b) Ru(DIPh)Macro, a metal complex constructed to
contain a central DNA-binding domain (Ru(DIP)32+) with two
tethered amine arms to chelate additional metal ions (Zn2+) to
deliver to the sugar-phosphate backbone and promote hydrolytic
strand cleavage.
(c) RNA site-specificially hydrolyzed by lead ion.
125
CHAPTER IV
Transition Metal Storage, Transport and
Biomineralization
Living organisms store and transport transition metals
both to provide appropriate concentrations of them for use in
metalloproteins or cofactors and to protect themselves against the
toxic effects of metal excesses; metalloproteins and metal
cofactors are found in plants, animals, and microorganisms.
In multicellular organisms, composed of a variety of
specialized cell types, the storage of transition metals and the
synthesis of the transporter molecules are not carried out by all
types of cells, but rather by specific cells that specialize in these
tasks. The form of the metals is always ionic, but the oxidation
state can vary, depending on biological needs. Transition metals
for which biological storage and transport are significant are, in
order of decreasing abundance in living organisms: iron, zinc,
copper, molybdenum, cobalt, chromium, vanadium, and nickel.
Iron is the most common transition metal in biology. 6,7
Its use has created a dependence that has survived the appearance
of dioxygen in the atmosphere ca. 2.5 billion years ago, and the
concomitant conversion of ferrous ion to ferric ion and insoluble
rust.
The average amount of iron in the human body (70 kg) is
ca. 5 g; iron is thus the most abundant transition metal in our
organism. About 70% of this amount is used for oxygen transport
and storage (haemoglobin, myoglobin), almost 30% are stored in
ferritins (iron storage proteins), and about 1% is bound to the
transport protein transferrin and to various irondependent
enzymes:
126
Haeme-type
(e.g.
cytochromes,
haemoglobin)
Iron-sulfur proteins
(e.g. ferridoxines,
Rieske proteins)
Two iron centers (e.g.
ribonucleotidereductase)
Three properties of iron can account for its extensive use
in terrestrial biological reactions:
(a) facile redox reactions of iron ions;
(b) an extensive repertoire of redox potentials
available by ligand substitution or modification;
(c) abundance and availability under conditions
apparently extant when terrestrial life began.
The processes and reactions in which iron participates are
crucial to the survival of terrestrial organisms, and include
ribonucleotide reduction (DNA synthesis), energy production
(respiration), energy conversion (photosynthesis), nitrogen
reduction, oxygen transport (respiration, muscle contraction), and
oxygenation (e.g., steroid synthesis, solubilization and
detoxification of aromatic compounds). On the other hand, Fe(III)
has a low solubility under physiological conditions ( 10-18 M),
requiring living organisms to adapt more efficient iron
storage/transport/usage mechanisms.
127
The Storage of Iron
Iron is stored mainly in the ferritins. Ferritin is a globular
protein complex consisting of 24 protein subunits (Figure 17, 18)
and is the primary intracellular iron-storage protein in both
prokaryotes and eukaryotes, keeping iron in a soluble and nontoxic form. Ferritin that is not combined with iron is called
apoferritin.
Figure 17. Structure of the murine ferritin complex
Figure 18. Crystallographic structure of mitochondrial ferritin
128
Ferritin is found in animals, plants, and even in bacteria;
the role of the stored iron varies, and includes intracellular use for
Fe-proteins or mineralization, long-term iron storage for other
cells, and detoxification of excess iron. Iron regulates the
synthesis of ferritin, with large amounts of ferritin associated with
iron excess, small or undetectable amounts associated with iron
deficiency.
The structure of ferritin is the most complete paradigm for
bioinorganic chemistry because of three features: the protein coat,
the iron-protein interface, and the iron core.
Protein Coat consists of twenty-four peptide chains (with
about 175 amino acids each), folded into ellipsoids, pack to form
the protein coat, which is a hollow sphere about 100 Å in
diameter; the organic surface is about 10 A thick (Figure 17).
Channels which occur in the protein coat at the trimer
interfaces may be involved in the movement of iron in and out of
the protein. Since the protein coat is stable with or without iron,
the center of the hollow sphere may be filled with solvent, with
Fe2O3 nH2O, or, more commonly, with both small aggregates of
iron and solvent. Very similar amino-acid sequences are found in
ferritin from animals and plants. Sorting out which amino acids
are needed to form the shape of the protein coat and the ligands
for iron core formation requires the continued dedication of
bioinorganic chemists; identification of tyrosine as an Fe(III)ligand adds a new perspective.
Iron-Protein Interface Formation of the iron core appears
to be initiated at an Fe-protein interface where Fe(II)-O-Fe(Ill)
dimers and small clusters of Fe(III) atoms have been detected
attached to the protein and bridged to each other by oxo/hydroxo
bridges.
Iron Core Only a small fraction of the iron atoms in
ferritin bind directly to the protein. The core contains the bulk of
the iron in a polynuclear aggregate with properties similar to
129
ferrihydrite, i.e. hydrous ferric oxide [Fe2O3(H2O)n] with various
amounts of phosphate. The overall composition of the iron
nucleus is 8FeO(OH)·FeO(H2PO4). Channels of threefold
symmetry and a width of 10 Å allow for an exchange of Fe3+
between the interior and exterior.
As many as 4,500 iron atoms can be reversibly stored
inside the protein coat in a complex that is soluble; iron
concentrations equivalent to 0.25 M [about 10 16-fold more
concentrated than Fe(III) ions] can be easily achieved in vitro.
Magnetite (Fe3O4) is another form of biological iron
derived, apparently, from the iron in ferritin. Magnetite plays a
role in the behavior of magnetic bacteria, bees, and homing
pigeons.
Ferritins – like many other proteins – exhibit high
symmetry. High symmetry (also found with higher organised
forms of life such as viruses, bacteria and even proteins in plants
and animals) makes less reactive – as a consequence of
minimised overall polarity – and thus has “a protective function”.
For some basic considerations on symmetry, also of relevance in
the context of the electronic configuration of metal ions in
coordination compounds
The Uptake and Transport of Iron
The extremely low solubility of the stored Fe(OH)3
[solubility product L = 2·10-39, solubility (pH 7) l =10-18 mol·l-1],
and thus the unavailability of iron in aqueous media under oxic
conditions, has forced many groups of organisms to develop
suitable systems for the mobilisation of iron. These systems, socalled siderophores excreted by the organisms, are multidentate
anionic ligands which form extremely stable complexes with Fe3+
(complex formation constants up to 1050 M-1). The functional
groups of these ligands are, in many cases, catecholates (o130
hydroxyquinolates), as in the case of enterobactin, or
hydroxamates (ferrioxamines and ferriochromes) (Figure 19).
The complexes are more or less globular, with the outer sphere
furnished with hydrophilic groups (amide and ester groups),
allowing for the water solubility and easy transport in the aquatic
medium. Internalization of the iron-loaded siderophore by the
organism typically takes place by endocytosis; the cytosolic
remobilisation of the iron either by reduction of Fe3+ to Fe2+ and
recycling of the siderophore, or by oxidative destruction of the
siderophore.
(a)
(b)
Figure 19. Structures of enterobactin (a) and ferrioxamine E (b)
131
Iron, when taken up with the food and processed in the
mouth (chewing, admixture of saliva) is mostly present in its
ferric (Fe3+) form and thus gets into the gastro-intestinal tract as
Fe3+. In case of an intact milieu in the small intestines, ferric iron
is reduced to its ferrous form (Fe2+). Only in this oxidation state
can iron be absorbed by the epithelium cells of the mucosa. For
transfer to the blood serum, reoxidation to Fe3+ is necessary. The
oxidation Fe2+ → Fe3+ + e- in the mucosa is catalyzed by a copper
enzyme (ceruloplasmin, containing 7 copper centres: Cu+ →
Cu2+). The Fe3+ ions are then taken up by apotransferrin (H2Tf);
simultaneously, carbonate is coordinated to iron. Fe3+-Tf is the
transport form for iron. The iron-loaded transferrin, (Figure 20)
delivers iron to sites of potential use (e.g. incorporation into
protoporphyrin IX and generation of haemoglobin), or stored in
iron storage proteins (ferritins).
The delivery of iron affords reduction from the ferric to
the ferrous state; a reductant employed here is ascorbate (vitamin
C):
uptake:
H2Tf + Fe3+ + HCO3- → [(Tf)FeIII(CO3)]- + 3H+
release:
[(Tf)FeIII(CO3)]- + e- + 3H+ → H2Tf + HCO3- + Fe2+
132
Figure 20. The FeIII – carbonate – transferring complex.
Biomineralization of Iron
Biomineralization is the process by which living
organisms produce minerals, often to harden or stiffen existing
tissues. Such tissues are called mineralized tissues.
Biologically formed iron ore is by far the most common
and thoroughly studied process of biomineralization There are
five major variations on iron biomineralization that incorporates
both BIM and BCM: Hydroxides and oxides, phosphates,
silicates, sulfates and sulfides.
Hydroxides and oxides. Iron oxides, as one of the beststudied classes of biominerals containing transition metals. Three
different forms of biological iron oxides appear to have distinct
133
relationships to the proteins, lipids, or carbohydrates associated
with their formation and with the degree of crystallinity.
Magnetite, on the one hand, often forms almost perfect
crystals inside lipid vesicles of magneto-bacteria. Ferrihydrite, on
the other hand, exists as large single crystals, or collections of
small crystals, inside the protein coat of ferritin; however, iron
oxides in some ferritins that have large amounts of phosphate are
very disordered. Finally, goethite [-FeO(OH)] and lepidocrocite
[-FeO(OH)] form as small single crystals in a complex matrix of
carbohydrate and protein in the teeth of some shellfish (limpets
and chitons); magnetite is also found in the lepidocrocitecontaining teeth.
The differences in the iron-oxide structures reflect
differences in some or all of the following conditions during
formation of the mineral: nature of co-precipitating ions, organic
substrates or organic boundaries, surface defects, inhibitors, pH,
and temperature. Magnetite can form in both lipid and
protein/carbohydrate environments, and can sometimes be
derived from amorphous or semicrystalline ferrihydrite-like
material (ferritin). However, the precise relationship between the
structure of the organic phase and that of the inorganic phase has
yet to be discovered.
Synthetic iron complexes have provided models for two
stages of ferritin iron storage and biomineralization: (1) the early
stages, when small numbers of clustered iron atoms are bound to
the ferritin protein coat, and (2) the final stages, where the bulk
iron is a mineral with relatively few contacts to the protein coat.
In addition, models have begun to be examined for the
microenvironment inside the protein coat (Figure 21).
134
Figure 21. The structure of a model for a possible intermediate in
the formation of the ferritin iron core. The complex consists of 11
Fe(III) atoms with internal oxo-bridges and a coat of benzoate
ligands; the Fe atoms define a twisted, pentacapped trigonal
prism.
Sulfides. Following ores that are the result of oxidization,
the sulfides, formed in anoxic environments are probably the
most important ores for mining. Pyrite is one of the most
common iron sulfides and has been connected in some cases to
bacterial activities. Sulfides are the end-products of bacterial
metabolism of the energy rich sulfates. The sulfide generated by
the bacteria reacts with iron oxides or hydroxides and form
135
monosulfide phases as FeS (and mackinawite (Fe,Ni)1-2S) but
also elemental sulfur. The latter then becomes the oxidant
required to convert FeS to pyrite FeS2.
Sulfates. As a result of oxidization in acidic conditions of
ferrihydrates and also iron sulfides bacteria can produce ferric
hydroxysulfates. The exact process of this in detail on a bacterial
level is still a matter of debate.
Phosphates. Phosphates are a common source of energy
for bacteria and occur in abundance in, for example anoxic water
environments. Phosphate is often dissolved from underlaying
apatite (Ca5(PO4)3(F,Cl,OH)) rich rocks or a product from the
decay of organic material. The formation of iron phosphate
minerals through microbes such as strengite (FePO42H2O),
which is a hydrated iron phosphate created by organic acids, has
been directly observed when analyzing biofilms growing on
phosphorite sediments. Associations with simple iron oxides
(FeO) and goethite has also been found in phosphorite sediments
from Paleozoic to Cenozoic times. These biofilms, or lichens,
which are symbiotical concentrations of bacteria and fungi
concentrate and immobilize metals from the phosphates.
Phosphorite sediments can sometime form banded layers of
hundred of meters in thickness with thinner layers of
phosphorous iron ores.
Silicates. Bacteria can also form iron rich silicates,
primarily found in acidic hot spring sediments. Examination of
such sediments has revealed bacterial cells completely encrusted
in these minerals. They are presumed to be formed as a result of
iron binding to anionic cells and where dissolved silica in the
spring then was subsequently added to the growing minerals in
136
the process. Another process in which silicates form has been
observed in biofilms in clays in freshwater environments. Here
(Fe, Al)-silicates similar to chamonite ((Fe5Al)(Si3Al)10(OH)8)
and kaolinite (Al4(Si4O10)(OH)4), but with a poor crystallinity was
found. These are therefore thought to have formed as a result of
reaction between the silica, metals and possibly metals within the
cell since it is well known that fresh water bacteria can bind and
immobilize metals. The process of forming a silicate is then
completed through diagenesis.
Conclusion
Transition play key roles in such biological processes. Among
them, Fe predominates in terrestial abundance; since Fe is
involved in a vast number of biologically important reactions, its
storage and transport have been studied extensively. Two types of
Fe carriers are known: specific proteins and low-molecularweight complexes. In higher animals, the transport protein
transferrin binds two Fe atoms with high affinity; in
microorganisms, iron is transported into cells complexed with
catecholates or hydroxamates called siderophores; and in plants,
small molecules such as citrate, and possibly plant siderophores,
carry Fe. Iron complexes enter cells through complicated paths
involving specific membrane sites (receptor proteins). A problem
yet to be solved is the form of iron transported in the cell after
release from transferrin or siderophores but before incorporation
into Fe-proteins.
The problems of storage, transport and biomineralization
of other transitional metals, such as Cu, Mo, Cr, Co, Mn, V and
Zn are proposed as research topics for the students and are not
discussed here.
137
CHAPTER V
Biological dioxygen transport system
The study of oxygen transfer in animals and plants
represents a large and diverse field of research, covering not only
different levels of organisation ranging from the whole organism
through to subcellular organelles, and different mechanisms such
as convection and simple or facilitated diffusion, but also, of
necessity, invoking diverse methodological approaches.
Most organisms require molecular oxygen in order to
survive. The dioxygen isused in a host of biochemical
transformations, although most is consumed in the reaction
O2 + 4H+ + 4e- 2H2O
that is the terminal (or primary) step of oxidative
phosphorylation.
An elegant three-component system has evolved to
transport dioxygen from regions of high abundance (water and
air) to regions of relatively low abundance and high demand (the
interior cells of the organism). This process is illustrated in
Figure 22.
138
Figure 22. Oxygen sequestration and transport in the
generalized organism.
The central component is a dioxygen-carrier protein
where the dioxygen-binding site, that is, the so-called "active
site," is a complex either of iron or of copper.
The second component of the dioxygen-transport system
facilitates the sequestration of dioxygen by the dioxygen-carrier
protein. Specialized organs, such as lungs in air-breathing
creatures or gills in fish, offer a very large surface area to the
outside environment in order to facilitate diffusion.
139
The third component is the delivery system. The oxygen
carrier is dissolved or suspended in a fluid, called blood plasma
or hemolymph, that is pumped throughout the animal by another
specialized organ, the heart, through a network of tubes, the
blood vessels.
Structure and characteristics of dioxygencarrier proteins
Hemoglobins are the most widely distributed family of
dioxygen carriers. Their active site consists of an iron porphyrin
(heme) group embedded in the protein. Hemoglobin is a globular
protein (i.e., folded into a compact, nearly spherical shape) and
consists of four subunits, as shown in Figure 23. Each protein
subunit is an individual molecule that joins to its neighboring
subunits through intermolecular interactions.
Figure 23. The molecular model of hemoglobin.
140
Each subunit contains a heme group, which is displayed
using the ball-and-stick representation in Figure 23. Each heme
group contains an iron atom that is able to bind to one oxygen
(O2) molecule. Therefore, each hemoglobin protein can bind four
oxygen molecules.
In the body, the iron in the heme is coordinated to the four
nitrogen atoms of the porphyrin and also to a nitrogen atom from
a histidine residue (one of the amino-acid residues in
hemoglobin) of the hemoglobin protein (Figure 24). The sixth
position (coordination site) around the iron of the heme is
occupied by O2 when the hemoglobin protein is oxygenated.
Figure 24. The coordination mode of iron in a
deoxygenated hemoglobin.
141
The heme group is nonplanar when it is not bound to
oxygen; the iron atom is pulled out of the plane of the porphyrin,
toward the histidine residue to which it is attached. This
nonplanar configuration is characteristic of the deoxygenated
heme group, and is commonly referred to as a "domed" shape.
The valence electrons in the atoms surrounding iron in the heme
group and the valence electrons in the histidine residue form
"clouds" of electron density. (Electron density refers to the
probability of finding an electron in a region of space.) Because
electrons repel one another, the regions occupied by the valence
electrons in the heme group and the histidine residue are pushed
apart. Hence, the porphyrin adopts the domed (nonplanar)
configuration and the Fe is out of the plane of the porphyrin ring
(Figure 25, left). However, when the Fe in the heme group binds
to an oxygen molecule, the porphyrin ring adopts a planar
configuration and hence the Fe lies in the plane of the porphyrin
ring (Figure 25, right).
Figure 25. Electron-density clouds of the deoxygenated
heme group (pink) and the attached histidine residue (light blue)
in deoxygenated (left) and oxygenated (right) hemoglobin
142
The shape change in the heme group has important
implications for the rest of the hemoglobin protein, as well. When
the iron atom moves into the porphyrin plane upon oxygenation,
the histidine residue to which the iron atom is attached is drawn
closer to the heme group. This movement of the histidine residue
then shifts the position of other amino acids that are near the
histidine (Figure 26). When the amino acids in a protein are
shifted in this manner (by the oxygenation of one of the heme
groups in the protein), the structure of the interfaces between the
four subunits is altered. Hence, when a single heme group in the
hemoglobin protein becomes oxygenated, the whole protein
changes its shape. In the new shape, it is easier for the other three
heme groups to become oxygenated. Thus, the binding of one
molecule of O2 to hemoglobin enhances the ability of hemoglobin
to bind more O2 molecules. This property of hemoglobin is
known as "cooperative binding."
Hemoglobin exists in two forms, a taut (tense) form (T)
and a relaxed form (R). Various factors such as low pH, high CO2
and high 2,3-Bisphosphoglyceric acid (2,3-Bisphosphoglycerate
or 2,3-BPG) at the level of the tissues favor the taut form, which
has low oxygen affinity and releases oxygen in the tissues.
Conversely, a high pH, low CO2, or low 2,3 BPG favors
the relaxed form which can better bind oxygen. The partial
pressure of the system also affects O2 affinity where, at high
partial pressures of oxygen, the tense state is favored. Inversely,
at low partial pressures, the relaxed state is favored. Additionally,
the binding of oxygen to the Iron-II heme pulls the iron into the
plane of the porphryn ring, causing a slight conformational shift.
The shift encourages other similar ligands to bind to the three
remaining hemes within hemoglobin (thus, oxygen binding is
cooperative).
143
Figure 26. Structures of deoxygenated (left) and
oxygenated (right) forms of hemoglobin
The structure of hemoglobins varies across species.
Hemoglobin occurs in all kingdoms of organisms, but not in all
organisms. Primitive species such as bacteria, protozoa, algae,
and plants often have single-globin hemoglobins. Many nematode
worms, molluscs, and crustaceans contain very large multisubunit
molecules, much larger than those in vertebrates. In particular,
chimeric hemoglobins found in fungi and giant annelids may
contain both globin and other types of proteins.
Among the other oxygen-binding proteins, we can mention
the following:
1.
Myoglobin (Figure 27) is found in the muscle
tissue of many vertebrates, including humans,
it gives muscle tissue a distinct red or dark gray
color. It is very similar to hemoglobin in
structure and sequence, but is not a tetramer;
instead, it is a monomer that lacks cooperative
144
binding. It is used to store oxygen rather than
transport it.
Figure 27. Example of molecular structure of myoglobin and
magnified view.
2.
Hemocyanin (Figure 28) is the second most
common oxygen-transporting protein. It is
found in the blood of many arthropods and
molluscs.
Figure 28. Deoxygenated (left) and oxygenated (right)
hemocyanin.
145
Oxygenation causes a color change between
the colorless Cu(I) deoxygenated form and the
blue Cu(II) oxygenated form. Hemocyanins
carry oxygen in the hemolymph of most
molluscs, and some arthropods. They are
second only to hemoglobin in frequency of use
as an oxygen transport molecule. Unlike the
hemoglobin in red blood cells found in
vertebrates, hemocyanins are not bound to
blood cells but are instead suspended directly
in the hemolymph.
3.
Hemerythrin (Figure 29) is an oligomeric
protein responsible for oxygen transport in the
marine invertebrate phyla of sipunculids,
priapulids, brachiopods, and in a single annelid
worm, magelona.
Figure 29. Active sites of hemerythrin before and after
oxygenation.
Hemerythrin does not, as the name might
suggest, contain a heme fragment and
represents a non-heme protein.
146
The mechanism of dioxygen binding is
unusual. Most O2 carriers operate via
formation of Dioxygen complexes, but
hemerythrin holds the O2 as a hydroperoxide.
The site that binds O2 consists of a pair of iron
centres. The iron atoms are bound to the
protein through the carboxylate side chains of
a glutamate and aspartates as well as through
five histidine residues. Hemerythrin and
myohemerythrin are often described according
to oxidation and ligation states of the iron
centre:
Fe2+—OH—Fe2+
Fe2+—OH—Fe3+
Fe3+—O—Fe3+—OOHFe3+—OH—Fe3+—R
(any other ligand)
deoxy (reduced)
semi-met
oxy (oxidized)
met (oxidized)
Deoxyhemerythrin contains two high-spin
ferrous ions bridged by hydroxyl group (A).
One iron is hexacoordinate and another is
pentacoordinate. A hydroxyl group serves as a
bridging ligand but also functions as a proton
donor to the O2 substrate. This proton-transfer
result in the formation of a single oxygen atom
(μ-oxo) bridge in oxy- and methemerythrin. O2
binds to the pentacoordinate Fe2+ centre at the
vacant coordination site (B). Then electrons
are transferred from the ferrous ions to
generate the binuclear ferric (Fe3+, Fe3+) centre
147
4.
with bound peroxide (C). It appears pink/violet
when oxygenated, clear when not.
Chlorocruorin is present in the blood plasma of
many annelids. It carries an abnormal heme
group structure (Figure 30). This enormous
macromolecule is typically found free floating
in the plasma, and not contained within red
blood cells.
Figure 30. Structure of the active center of chlorocruorin
5.
148
The affinity of chlorocruorin for oxygen is
weaker than that of most haemoglobins. It
appears green when deoxygenated and red
when oxygenated.
Vanabins are also known as vanadium
chromagens, they are found in the blood of sea
squirts. There were once hypothesized to use
the rare metal vanadium as an oxygen binding
6.
prosthetic group. However, although they do
contain vanadium by preference, they
apparently bind little oxygen, and thus have
some other function, which has not been
elucidated.
Erythrocruorin is found in many annelids,
including earthworms, it is a giant free-floating
blood protein containing many dozens –
possibly hundreds – of iron- and heme-bearing
protein subunits bound together into a single
protein complex with a molecular mass greater
than 3.5 million daltons.
Requirements for Effective Oxygen Carriers
In order for dioxygen transport to be more efficient than
simple diffusion through cell membranes and fluids, it is not
sufficient that a metalloprotein merely binds dioxygen. Not only
is there an optimal affinity of the carrier for dioxygen, but also,
and more importantly, the carrier must bind and release dioxygen
at a rapid rate. These thermodynamic and kinetic aspects are
illustrated in Figure 31 for the process
where M is an oxygen carrier, for example hemocyanin.
Thermodynamic or equilibrium aspects are summarized
by G in Figure 31.
149
Figure 31. Schematic diagram of energy changes in dioxygen
binding.
As it is illustrated, G is negative, and thus the forward
reaction, dioxygen binding, is spontaneous. The equilibrium
constant (K) is given by
where a is activity of species or in terms of concrntrations and
partial pressure of oxygen:
150
It is very convenient to express the affinity as the partial
pressure of dioxygen required for half-saturation of the species
M, Pl/2(O2). Under such conditions, [M] = [MO2], one obtains
The dioxygen affinity is composed of enthalpic (H) and
entropic (S) components, with
Within a family of oxygen carriers the values of ASo and
Mfo are usually similar. Large deviations (such as a change of
sign) are therefore indicative of a change in the nature of the
oxygen-binding process.
Non-cooperative Dioxygen Binding
If the oxygen-binding sites M are mutually independent
and noninteracting, as in moderately dilute solutions of
monomeric molecules, then the concentration of species MO2 as a
function of the partial pressure of O2 is generally well fit by a
Langmuir isotherm with the fractional saturation of dioxygen
binding sites,  as
Such binding, where the dioxygen sites are independent of
each other, is termed non-cooperative.
151
Cooperative Dioxygen Binding
Many dioxygen-binding proteins are not independent
monomers, with only one dioxygen-binding site, but oligomeric
species with the protein comprising two or more similar subunits.
The subunits may be held together by van der Waals' forces or by
stronger interactions, such as hydrogen bonds or salt bridges, or
even by covalent bonds. For example, most mammalian
hemoglobins are tetramers where either none, one, two, three, or
all four sites may be occupied by dioxygen (Figure 32).
Thus, the binding or release of dioxygen at one site may
affect the affinity and kinetics of ligand binding and release at a
neighboring site. As a result, the saturation curve becomes
sigmoidal in shape (Figure 33). When cooperativity is positive,
the affinity of a vacant site is increased by occupancy of an
adjacent one.
152
Figure 32. Diagram of tetrameric hemoglobin, showing statistical
weights of different saturations
153
Figure 33. Binding curves of cooperative and non-cooperative
binding of dioxygen.
Dioxygen reactions
Molecular dioxygen in its ground state is shown below
(Figure 34). The bond order for this configuration is 2, containg
one  and one  bond. Hund's rule of multiplicity dictates that the
two electrons in the 2p* orbitals occupy different degenerate
orbitals. Since they are in different orbitals they will have parallel
spins and a net spin angular momentum (that will be in a triplet
state).
154
Figure 34. MO energy diagram for a dioxygen molecule.
Chemical reactions forming singlet molecules from triplet
and singlet reactants are forbidden by Wigner's spin selection
rule. Therefore the number of unpaired electrons must remain the
same before and after each elementary step of a chemical
reaction. For these reasons, is impossible for the following
reaction to go in one fast, concerted step:
½ 3O2 + 1X  1XO
 

155
The arrows represent electron spins:  represents a
singlet molecule with all electron spins paired;  represents a
triplet molecule with two unpaired electrons; and  represents a
doublet molecule, also referred to as a free radical, with one
unpaired electron.
Majority of the organic compounds are characterized by
co-valent bonds where all the electrons are pared (singlet state).
Thus, the triplet multiplicity is the actual reason, why most
reactions of oxygen with organic substances, although being
exergonic, do not proceed at room temperature. It is said that
reactions of organic compunds with oxygen are kinetically
inhibited. This effect enables our life in an oxygen containig
atmosphere.
The pathways that do not violate the spin restriction are
all costly in energy, resulting in high activation barriers. For
example, one can suppose the formation of the excited triplet
state of the oxygenated organic product which is followed by a
slow spin conversion to a singlet product:
½ 3O2 + 1X  3XO
 

3
XO  1XO (slowly)


Another pathway in which O2 is excited to a singlet state
that then reacts with the substrate would be spin-allowed but
require a high activation energy:
O2 + 22.5 kcal/mol  1O2


3
156
½ 1O2 + 1X  1XO
  
One pathway for a direct reaction of triplet ground-state
dioxygen with a singlet ground-state organic substrate that can
occur readily without a catalyst begins with the one-electron
oxidation of the substrate by dioxygen. The products of such a
reaction would be two doublets, i.e., superoxide and the one
electron oxidized substrate, each having one unpaired electron.
These free radicals can diffuse apart and then recombine with
their spins paired. Such a mechanism has been shown to occur for
the reaction of dioxygen with reduced flavins:
However, this pathway requires that the substrate be able
to reduce dioxygen to superoxide, a reaction that requires an
157
unusually strong reducing agent (such as a reduced flavin), since
dioxygen is not a particularly strong one-electron oxidizing agent.
The result of these kinetic barriers to dioxygen reactions
with most organic molecules is that uncatalyzed reactions of this
type are usually quite slow.
Most biological conversions involving dioxygen require
enzymatic catalysis. The functions of the metalloenzymes for
which dioxygen is a substrate are, therefore, to overcome the
kinetic barriers imposed by spin restrictions or unfavorable oneelectron reduction pathways, and, for the oxygenase enzymes, to
direct the reactions and make them highly specific.
Transition metals in the correct oxidation state can react
directly with triplet O2 to form dioxygen adducts that can
participate in reaction pathways leading to the incorporation of
oxygen into organic substrates. Dioxygen, in this case, is not only
coordinated, but also activated and made available to the
substrate.
In binding to metals, O2 effectively functions both as a acid, accepting into its *-orbitals electron density from the filled
d-orbitals of the metal, and as a -donor, donating electron
density into an empty metal d-orbital (Figure 35).
Figure 35. Types of interaction of the electron density of O2
with d-orbitals of iron.
158
The metal d-orbitals, now having partial porphyrin *
character, are split, as shown in Figure 36.
Figure 36. d-Orbital splitting of Fe3+ ions in metalloporphyrins as
a function of number and ligand-field strength.
Iron(II) tends to exist in a high-spin configuration where
unpaired electrons exist in Eg antibonding orbitals. Iron(III) has
an odd number of electrons, and thus must have one or more
unpaired electrons, in any energy state.
All of these known structures are paramagnetic (have
unpaired electrons), not diamagnetic. Thus, a non-intuitive (e.g., a
higher-energy for at least one species) distribution of electrons in
159
the combination of iron and oxygen must exist, in order to
explain the observed diamagnetism and no unpaired electrons.
The three logical possibilities to produce diamagnetic (no
net spin) Hb-O2 are:



160
Low-spin Fe2+ binds to singlet oxygen. Both lowspin iron and singlet oxygen are diamagnetic.
However, the singlet form of oxygen is the higherenergy form of the molecule.
Low-spin Fe3+ binds to O2- (the superoxide ion)
and the two unpaired electrons couple
antiferromagnetically,
giving
diamagnetic
properties.
Low-spin Fe4+ binds to peroxide, O22-. Both are
diamagnetic.
CHAPTER VI
Bioinorganic Chemistry of dinitrogen
fixation
Nitrogen is a critical limiting element for plant growth
and production. It is a major component of chlorophyll, the most
important pigment needed for photosynthesis, as well as amino
acids, the key building blocks of proteins. It is also found in other
important biomolecules, such as ATP and nucleic acids. Even
though it is one of the most abundant elements (predominately in
the form of nitrogen gas (N2) in the Earth’s atmosphere), plants
can only utilize reduced forms of this element. Plants acquire
these forms of “combined” nitrogen by: 1) the addition of
ammonia and/or nitrate fertilizer (from the Haber-Bosch process)
or manure to soil, 2) the release of these compounds during
organic matter decomposition, 3) the conversion of atmospheric
nitrogen into the compounds by natural processes, such as
lightning, and 4) biological nitrogen fixation
Nitrogen fixation is the biogenic and non-biogenic
transformation of elemental N2 into nitrogen compounds (Figure
37), affording to overcome the bonding energy between the two
trebly bonded nitrogen atoms (949 kJ/mol). Nitrogen fixation is a
key reaction of the biological nitrogen cycle. The biogenic
fixation, carried out by free living nitrogen-fixing bacteria
(Azotobacter) and cyanobacteria (“blue-green algae”, Anabaena),
some archaea, and by symbiotic bacteria associated with
leguminous plants (Rhizobium), leads to ammonium ions.
Biogenic fixation accounts for about 60% of the overall nitrogen
supply.
161
Figure 37. The nitrogen cycle.
The N2 molecule has a triple bond with energy 225
kcal/mole. The challenge to which nitrogenase rises is to break
and reduce at a reasonable rate the extremely strong N≡N triple
bond. The kinetic inertness of N2 is highlighted by the fact that
carrying out reactions "under nitrogen" is considered equivalent
to doing the chemistry in an inert atmosphere. Despite this kinetic
inertness, thermodynamically the reduction of N2 by H2 is a
favorable process,
and at pH 7 the reaction
162
has an E0 value that makes it easily accessible to biological
reductants such as the low-potential ferredoxins.
The thermodynamically favorable reduction of N2 to
2NH3 is a six-electron process. Unless a concerted 6e , 6H +
process can be effected, intermediates between N2 and NH3 must
be formed. However, all the intermediates on the pathway
between N2 and NH3 are higher in energy than either the reactants
or the products (Figure 38).
Figure 38. Energetics of N2, NH3, and some potential
intermediates along the reaction pathway for their
interconversion.
Several factors may allow this barrier to be overcome.
First, the six-electron reduction might be carried out in a
163
concerted or near-concerted manner to avoid the intermediates
completely. Alternatively, the intermediates could be complexed
at metal centers to stabilize them to a greater extent than either
the reactants or products. Finally, the formation reaction for the
unfavorable intermedi ate could be coupled with ATP hydrolysis
or with the evolution of dihydrogen, each a favorable process, so
that the overall process is favorable.
The Mo-nitrogenases
The action of the Mo-nitrogenase enzyme involves the
functioning of two separately isolatable component proteins, as
sketched in Figure 39.
Figure 39. Organisation of nitrogenase (top), and the
structure of the M cluster (bottom).
The electrons necessary for the reduction of dinitrogen are
delivered by an iron protein containing a cubanelike [4Fe,4S]
ferredoxin. Primary e- acceptor is the P cluster of the FeMoco,
164
the ironmolybdenum-cofactor. Two ATP (activated by Mg2+) are
hydrolysed per electron transferred. The FeMoco contains two P
and two M clusters, arranged in such a way that the complete
cofactor attains C2 symmetry. The P cluster is a double cubane
containing the Fe8S7 core. The reduction equivalents are then
further transported to the M cluster, a Fe7MoS9 core, which is
responsible for the final activation and reduction of N2. The cage
formed by the metal centres of the M cluster contains electron
density which can be interpreted in terms of a nitrogen atom.
The M cluster is connected to the protein matrix by just
one Cys and a His, the latter coordinated to Mo. The coordination
environment of Mo is supplemented by the vicinal hydroxide and
carboxylate of homocitrate. In which way activation and
reduction of N2 takes place is unknown. The properties of some
representative nitrogenases are arranged in the Table 5.
Table 5. Properties of some representative nitrogenases.
165
The V-nitrogenases
In the case of insufficient molybdenum supply, or at low
temperatures, a vanadium-nitrogenase is activated (which is more
efficient at lower temperatures than the Mo version). The isolated
enzyme was reported to be similar to the Mo enzyme, but had a
lower activity and an altered substrate specificity.
One of the two components of the V-nitrogenase system
is extremely similar to the Fe protein of nitrogenase. Both Fe
proteins have an 2 subunit structure, and contain a single Fe4S4
cluster.
A major difference between the V and Mo enzymes lies in
substrate specificity and product formation. the FeV nitrogenase
has a much lower reactivity toward acetylene than does the Mo
system. Furthermore, whereas the FeMo system exclusively
produces ethylene from acetylene, the FeV system yields
significant amounts of the four-electron reduction product,
ethane.
The all-iron nitrogenase
The first sign that there is yet another alternative
nitrogenase again came from genetic studies. The finding of the
all-iron nitrogenase, if fully confirmed, will add significantly to
the comparative biochemistry of nitrogen fixation. Speculatively,
one might suggest that the concomitant absence of V and Mo
suggests that nitrogen fixation need not directly involve the noniron heterometal in the cofactor cluster.
166
Model systems
Three types of model systems for nitrogenase may be
considered. First, there are transition-metal sulfide clusters that
resemble the FeMoco or FeVco centers of the active proteins.
Both X-Ray structural analysis on single crystals, and quantumchemical modeling of the enzyme active sites are used in order to
predict the possible structures of intermediate complexes (Figure
40).
Figure 40. Dinitrogen bound intermediate of the active
site of nitrogenase from DFT calculations.
A second approach uses the reactions of N2 and related
substrates or intermediates with metal centers in order to gain
insights into the way in which transition-metal systems bind N2
and activate it toward reduction. As an example, in 2011
Arashiba et al. reported yet another system with a catalyst again
based on molybdenum but with a diphosphorus pincer ligand:
167
And finally, there are other inorganic systems that display
some of the structural and possibly some of the reactivity
characteristics of the nitrogenase active sites without binding or
reducing N2 or precisely mimicking the active center.
168
EXPERIMENTS
In Bioinorganic Chemistry
169
170
EXPERIMENT 1
Electronic structures of atoms and ions. Molecular
diagrams of molecules and molecular ions. Bond
orders and multiplicities of electronic states
Goals. To understand how electronic configurations of
atoms and molecules affects their possible valence states and
configurations of reactional centers of biomolecular complexes.
Theoretical part.
In atomic physics and quantum chemistry, the electronic
configuration is the distribution of electrons of an atom or
molecule (or other physical structure) in atomic or molecular
orbitals. According to the laws of quantum mechanics, an energy
is associated with each electron configuration and, upon certain
conditions, electrons are able to move from one orbital to another.
Knowledge of the electronic configuration of different
atoms and molecules is useful for describing the chemical bonds
of different classes of chemical compounds.
Experimental part.
1. Draw the electronic configurations of the following atoms and
ions: Zn and Zn2+; Cu, Cu+ and Cu2+; Co, Co2+ and Co3+; Fe,
Fe2+, Fe3+ and Fe4+; Mo, Mo2+ and Mo6+; V, V2+, V3+ and V5+.
For each group compare the relative stabilities of species.
Indicate the number of unpaired electrons.
2. Draw the electronic configurations of phosphorus, sulfur and
oxygen in the ground and possible excited states. Predict
171
possible valence states of the above elements and present the
formulae of the corresponding compounds.
3. Using the VB method, describe the structures of the following
molecules and ions: H2O, H3O+, NH3, NH4+. Indicate the type
of hybridization of atoms and their spatial disposition.
4. Explain in terms of the VB theory the following observations:
(a) CCl4 and SiCl4 are both tetrahedral;
(b) CO2 is linear but SO2 is angular;
(c) HSH angle in H2S is closer to 90 than the HOH angle in
H2O;
(d) PCl5 exists but not NCl5;
(e) BF3 is planar but NH3 is not.
5. The valence electron configuration of Hg is 6s2. What kind of
hybrid orbitals can be used to account for the linear geometry
of HgCl2?
6. Use the MO energy level diagrams to determine bond orders
and multiplicities of the ground states of the following
molecules and ions: O2, O2-, O22-, N2, N22-.
7. Which of the following pairs of molecules would you expect
to have the higher bond energy: (a) F2 and F2+; (b) NO and
NO-; (c) BN and BO; (d) NF and NO?
8. Explain in terms of the MO theory the following observations:
(a) the first ionization energy of NO is less than that of CO;
(b) the first ionization energy of O2 is less than that of the oxygen
atom.
172
EXPERIMENT 2
Electronic and spatial structures and chemical
properties of coordinate compounds
Goals. To study the reactions of synthesis, decomposition,
and exchange in the inner spheres of coordinate compounds. To
correlate colors of the coordinate compounds of the transitional
elements with the strengths of the ligand field.
Theoretical part.
A coordination complex or metal complex, consists of an
atom or ion (usually metallic), and a surrounding array of bound
molecules or anions, that are in turn known as ligands or
complexing agents.
Many of the properties of metal complexes are dictated by
their electronic structures. The electronic structure can be
described by a relatively ionic model that ascribes formal charges
to the metals and ligands. This approach is the essence of crystal
field theory (CFT). Crystal field theory, introduced by Hans
Bethe in 1929, gives a quantum mechanically based attempt at
understanding complexes.
More sophisticated models embrace covalency, and this
approach is described by ligand field theory (LFT) and Molecular
orbital theory (MO). They can handle a broader range of
complexes and can explain complexes in which the interactions
are covalent.
173
Experimental part.
I. Formation of complex compounds.
1. Put 4-5 drops of the aqueous solution of copper chloride into
a test tube. Add ammonium hydroxide until the precipitate is
formed. Add excess of NH4OH and observe the formation of
a complex compound. Indicate the change in color while
transformation of an aqua-complex of copper(II) into the
ammine complex. Draw the electronic formula of the Cu2+
cation. Using the VB method, determine the type of
hybridization of the central atom in the ammine complex.
Draw the MO energy level diagrams for the squaric Cu(II)
complexes. Basing on the colors of the complexes, compare
the ligand field strengths of the water and ammonia ligands.
2. Put 4-5 drops of the aqueous solution of nickel chloride into a
test tube. Add Add 1 ml of the dimethylglyoxime solution and
observe the formation of a complex compound. Indicate the
change in color while transformation of an aqua-complex of
nickel(II) into the dimethylglyoximato-complex. Draw the
electronic formula of the Ni2+ cation. Draw the MO energy
level diagrams for the octahedric Ni(II) complexes. Basing on
the colors of the complexes, compare the ligand field
strengths of the water and dimethylglyoxime ligands.
II. Destruction of complex compounds.
1.Take the solution of chloride tetraammine copper(II), obtained
from the previous experiment, add dropwise diluted (1:1) nitric
acid until the complex compound is destroyed. Indicate the
change in color. Write the reaction in a net ionic form.
174
2.Take from 5 to 6 drops of CdCl2 and HgSO4 into sepaate test
tubes. Add dropwise a saturated solution of Na2SO3. Observe
the formation of precipitates of the corresponding sulfites and
their further transformation into complex compounds of the
composition of Na2[M(SO3)2], M= Cd2+ or Hg2+. Add the
solution of NaOH to both the solutions of complex compounds
and notice your observatins. Compare the instability constants
of Na2[Cd(SO3)2] and Na2[Hg(SO3)2] with the solubility
product constants of the corresponding metal hydroxides and
conclude about the possibility of the metal complexes to be
destroyed by the solution of NaOH.
III. Exchange of ligands in complex compounds.
1.Take 5 to 6 drops of the solution of FeCl3 in a test tube and add
2 to 3 drops of the solution of NH4SCN. Observe the change in
color while complex formation. Then add some crystals of
NH4F and shake well the test tube. Write down your
observations. Compare the instability constants of the complex
ions and conclude about the possibility of the reaction of ligand
exchange. Draw the electronic structures and determine the
types of hybridization if a ferric cation in the complexes of the
ligands of a weak and a strong field.
2.Perform the corresponding calculations and prove
experimentally that SCN- ligands can not replace fluoride
anions from the inner sphere of the ferric complex.
175
EXPERIMENT 3
Redox-properties of complexes of transition
metals
Goals. To study the redox-activities of central ions in
coordinate compounds.
Theoretical part.
Complex compounds may possess redox-properties and
be involved in the reactions where the oxidation state of the
central ion changes.
Two types of redox reactions of complex compounds are
possible.
i) The oxidation state changes without decomposition of
the inner sphere:
2K3[Fe3+(CN)6] + 2KI = 2K4[Fe2+CN)6] + I2
ii) The change in the oxidation state of the central atom
provokes the destruction of a complex compound:
Zn + 2K[Au+(CN)2] = 2Auo + K2[Zn(CN)4]
Experimental part.
I. Oxidation of the central ion.
1. Take 1 ml of the solution of KMnO4 in a test tube, add an
equal volume of a diluted solution of H2SO4 and then
drpowise K4[Fe(CN)6]. Observe the change in color of the
solution. Prove the formation of K3[Fe(CN)6] (probe with
FeSO4 solution). If necessary, peform test reactions of
K4[Fe(CN)6] and K3[Fe(CN)6] with the solutions of Fe(II) and
176
Fe(III) salts. Write and balance the reaction of oxidation of
K4[Fe(CN)6] into K3[Fe(CN)6] by KMnO4 in the acidic
medium.
2. Take 1 ml of the solution of CoCl2 and 2 ml of ammonia
solution in one test tube. Indicate the color of the ammine
complex of Co(II). Add 1 ml of a NH4Cl solution and 2 ml of
H2O2. Heat the test tube and indicate the change in color
because of oxidation of Co(II) into Co(III). Add H2O2 to the
neutral solution of CoCl2 in another test tube (do not add NH3
solution). Heat the solution and compare oxidating abilities of
Co2+ cations in [Co(H2O)6]2+ and [Co(NH3)6]2+ complexes.
Write and balance the redox-reaction.
II. Reduction of the central ion.
1. Take 1 ml of the solution of CuSO4 in a test tube and add
dropwise 25% solution of NH3 until the formation of a darkblue solution of [Cu(NH3)4]SO4. Put a zinc granule into the
solution and observe the changes on the surface of the
granule. Write and balance all the the reactions.
2. Mix equal volumes of AgNO3 and NaCl solutions (10 to 12
drops each) in a test tube. Dissolve the formed precipitate in
the concentrated ammonia solution. Add the Zn granule to the
solution. Write down and explain your observations. Write
and balance all the the reactions.
III. Redox reactions while complex formation.
Take 5 to 6 drops of any Co(II) salt solution into a test tube
and add 5 drops of CH3COOH and the same amount of KNO2.
Observe the formation of a yellow precipitate of K3[Co(NO2)6]. Is
the studied reaction a redox one? Write and balance it.
177
EXPERIMENT 4
Quantum-chemical modeling of tautomeric
forms of nucleic acids and their complexes
with metals
Goals. Application of methods of quantum chemical
modeling for prediction of tautomeric isomerism of organic
species and their complexes with metals.
Theoretical part.
Computational Methods for Bioinorganic Systems are
widely used. Proteins contain many atoms, too many to all be
treated using accurate electronic structure methods based on
quantum mechanics (QM) using current methods and computers.
As a result, people have developed hybrid "QM/MM" methods in
which the system is partitioned into a reactive "core" which is
treated using QM, and a remaining "environment" which is
treated using the much less expensive molecular mechanics
(MM). A variety of such QM/MM methods, depending on the
type of QM and MM used, and the way in which the interaction
of the QM and MM regions is treated. Potential energy surfaces
for proteins are also extremely complicated, with many similar
minima differing in the orientation of amino acid sidechains,
water molecules, etc.
For the nowdays, a large variety of computer packages of
programs for quantum chemical modeling purposes exist. The
mostly used are MOPAC, ChemOffice, HyperChem, Gamess and
Gaussian.
A semiempirical PM3 method is proposed for calculation
of relative stabilities of tautomers and conformers of organic
178
molecules and their complexes with metals. All the calculations
should be performed with the Gamess complex. The ChemOffice
package is to be used for construction and redaction of structures,
and for the visualization of the results the ChemCraft program is
proposed.
Experimental part.
1.Chose the object of investigation out of the proposed structures
(nucleic bases or amino acids).
2.Using the ChemOffice package, draw all possible tautomeric
forms of the molecule to be studied.
3.Optimize geometry using molecular mechanics approach and
then perform quantum chemical geometry optimization.
4.Save the coordinates of optimized structure into a file; show the
optimized bondlengths and valence angles using preferred
visualisation software (e.g. ChemCraft). Write down the heat of
formation and the total energy of the obtained conformer.
5.Repeat the procedure presented in points 3 and 4 for the other
possible tautomeric forms of the compound.
6.Basing on the calculated H or Etotal values of the tautomers,
conclude on their relative stabilities. Compare the calculated
results with the literary data on the form of existence of the
studied substance in crystalline form and in solutions.
7.Construct all the possible structures of the complexes of the
studied molecule with one of the metals (consult your
instructor). Pay attention to monodental or chelating
coordination and all possible active sites of the organic species.
8.Repeat the procedure presented in points 3 – 6. Make your
conclusion on the most probable structure of the complex
compound.
179
EXPERIMENT 5
Determination of ionization constants of amino acids
by potentiometry
Goals. Application of physico chemical methods
(potentiometry) for determination of ionization constants of organic
species.
Theoretical part.
Potentiometry involves measuring an electrical potential that is
related to a component in which someone is interested. For example,
many communities fluoridate water supplies to enhance the durability
of tooth enamel. One convenient way to measure fluoride concentration
is to use an electrode where the potential depends upon the
concentration of fluoride. Potentiometric electrodes are usually fast,
portable, and do not require extensive training to operate.
The
most
common
electrode in use is the glass
membrane pH electrode (see
figure). The difference in electrical
charge across the glass membrane
is the membrane potential that
depends only on the concentration
of H3O+ in the outer solution.
Organic acids usually
possess weak properties, i.a. their
dissociation is not full and
reversible; a dynamic equilibrium
exists, which is represented by the
following equation:
HA + H2O  H3O+ + A–
The equilibrium expression for this reaction is:
180
Ka 
 [ HA] 
[ H 3O  ][ A  ]
or [ H 3O  ]  K a   
[ HA]
 [A ] 
(1),
where Ka is the acid-ionization constant for the weak acid. Let us
assume that the initial dissociation of the weak acid is negligible. The
progressive addition of NaOH during the titration decreases the
concentration of HA and increases the concentration of its salt, NaA:
HA (aq) + NaOH (aq)  H2O(l) + NaA (aq)
The presence of both HA and its salt, NaA, creates a buffer
system, which resists a large change in pH. The ratio of [HA]/[A–]
changes only slightly; therefore, according to (1), the change in [H3O+]
(or pH) must also be small. The pH increases slowly until the
equivalence point is approached:
At the halfway point in the titration, exactly half of the HA
originally present will have been neutralized, and therefore the
concentrations of HA and A– will be equal.
[ H 3O  ]1 / 2 [ A  ]1 / 2
;
Ka 
[ HA]1 / 2
K a  [ H 3O  ]1 / 2 (2)
181
Thus, the ionization constant of a weak acid is equal to the
hydronium ion concentration at the halfway point in the titration; pKa =
pH1/2. This relationship is valid only if the initial dissociation of the acid
is negligible. When the degree of dissociation is appreciable, as in the
case of a very dilute solution, the pH at the midpoint of the titration
bears no relation to the value of Ka.
Amino acids are a little different from the other weak acids.
When the amino acids are placed in a strong acid like HCl, as will be
the case for this experiment, the amino acid will be completely
protonated and have a net ionic charge of +1. As the base is added in
the titration, the proton on the carboxylic acid will be removed first, so
that around a pH of 7, the zwitterionic form will exist. At the end of the
titration, the amino acid will be completely deprotonated.and have a
charge of -1:
pH 1 Net charge +1
pH 7 Net charge 0
O
R
pH 13 Net charge -1
O
O
OH
NH3+
Cationic form
R
O-
NH3+
Zwitterionic form
R
OH
NH2
Anionic form
A titration curve of aminoacids starting from strongly acidic
solutions therefore looks like:
182
Experimental part.
1. Prepare the necessary reagents and equipment:
Chemicals
Equipment
1 M HCl
Weighing boats
0.1 N NaOH
Spatula
Amino acid (assigned to each
student)
pH 4 and 7 buffers
Electronic pan balance
1-calibrated pH Checker
Ring stand
Buret and clamp
125 ml Erlenmeyer flask
2. Mass out a sample of your amino acid (not more than 0.1000 g) and
record the name of the amino acid and its mass.
3. Pour the sample into the pre-labeled Erlenmeyer flask and dissolve
the amino acid in 25 ml of water.
4. Obtain your pH checker and turn it on. Insert it into the amino acid
solution and allow it to stabilize. Once it has stabilized, add enough
1 M HCl to it to adjust the pH of the solution to a pH of about 1 to
1.5. Record that pH and the volume of HCl added.
5. Obtain and clean a 50 ml buret. Condition the buret with a
standardized sodium hydroxide solution of concentration
approximately 0.1 M. Place the buret in a buret holder on a ring
stand and fill with the sodium hydroxide solution. Make sure that
there are no bubbles neither in the buret nor in its tip. Record the
initial volume reading in your notebook showing two decimal
places.
6. Add stepwise 0.5 ml of the NaOH solution to the studied solution
while stirring, and record pH after each addition (in total 50 ml of
NaOH should be added). Arrange your results as a table:
V(NaO 0 0. 1. 1. 2. … … … … … … …
H), ml
5
0 5
0
pH
183
7. Using the results of the titration, set up the titration curve (pH vs.
V(NaOH)) and first-derivative curve (pH/V vs. V(NaOH)) (see
the figure above).
8. For pKCOOH determination, find the first endpoint volume (from
your first derivative curve) and divide that in half. Record the
experimental pKCOOH value.
9. For pKNH2 determination, find the first and second endpoint
volumes from your first derivative curve. Find the halfway volume
between the two. Back determine the pKNH2 at that volume from
your titration curve. Record the experimental pKNH2 value.
10. Insdead of points 7 – 9, you can also use a Hyperquad computer
program for calculations.
11. Search the pKCOOH and pKNH2 values in the internet databases and
calculate the error of your experiment.
184
EXPERIMENT 6
Determination of stability constants of
complexes of transitional metals by potentiometry
Goals. Application of physico chemical methods
(potentiometry) for determination of stability constants of
complexes of metals with organic species.
Theoretical part.
General approach.
Metal ions in solutions most often exist as solvates
(hydrates in aqueous solutions). The reaction of complex
formation may be considered as the reaction of ligand exchange:
М(Н2O)n + L → M(H2O)n-1,L + Н2O
where М is a metal ion, and L is a ligand.
During the complex formation, the coordinated (hydrated)
water molecules are consistently replaced by ions or molecules of
ligands to form MLn complex (n here indicates number of ligand
molecules). For easier presentation, the solvated metallic ion can
be presented in a “free” form (Мe+x). The reaction of complex
formation can be presented as:
mM+x + nL→MmLn
The thermodynamic constant of this reaction can be
expressed as following:
where f are activity coefficients.
185
In the case when the ionic strength I lies in the interval 0.1
– 0.5, the concentrational overall stability constant may be used:
The complex formation usually occurs stepwise:
M + L↔ML1
ML1 + L↔ML2
……………………..
……………………...
MLn-1 + L ↔ MLn
Stepwise stability constants are determing as following
equations:
The product of stepwise stability constant is known as an
overall formation constant:
β = К1·К2·...Кn
Potentiometric determinations.
A lot of methods of determination of stability constants of
complex compounds exist, and the method of potentiometric
titration is one of them. In this method, the ligand possessing
acidic properties is titrated by an alkali solution individually and
then in its mixture with a metal salt. As hydrogen ions of the
ligand (weak acid) are replaced by a metal according to the
equation:
M + xHL↔MLx + xH+,
the titration curve of the metal-to-ligand mixture will lie in a
lower pH range than the one of the individual ligand:
186
This difference in titration curves gives the possibility to
calculate the formation constant of a comples using
potentiometric titration and a special characteristic of the
equilibrium process of complex formation which is called
_
formation function, n :
C ( HL )  [ L ]
n
,
C (M )
_
where C(HL) an C(M) are total ligand and metal concentrations,
and [L-] is the equilibrium concentration of the non-complexed
ligand.
The formation function indicates the mean ligand
molecules bound to one metal ion. For a chelating ligand (i.e.
aminoacid):
log([L-]) = log(C(HL) – C(NaOH) – [H+] + [OH-]) – logP
[ H  ] 2[ H  ] 2
P

where K2 > K1.
K2
K1 K 2
187
In the case of formation of a 1 : 2 complexes of metals
with amino acids
_
n
2C ( NaOH )
C ( HL )
The total concentration of the amino acid includes
deprotonated, neutral, protonated, and coordinated species:
C(HL) = [L-] + [HL] + [H2L+] + [ML] + 2[ML2];
_
[ ML ]  2[ ML2 ]
n
;
C (M )
_
C(HL) = [L-] + [HL] + [H2L+] + n C(M);
and finaly,
_
_
K1 
n
_
;

(1  n) [ L ]
K2 
n 1
_
;
 = K1K2.

( 2  n) [ L ]
Experimental part.
1. Prepare the necessary reagents and equipment:
Chemicals
Equipment
1 M HCl
Weighing boats
0.1 N NaOH
Spatula
Amino acid (assigned to each
Electronic pan balance
student)
Metal salt (assigned to each
Buret and clamp
student)
pH 4 and 7 buffers
Ring stand
1-calibrated pH Checker
125 ml Erlenmeyer flask
188
2. Mass out a sample of your amino acid (the same name and the
same mass as in the previous experiment) and record the
name of the amino acid and its mass. Calculate the number of
moles of the sample.
3. Considering that a ML2 complex is formed, calculate the
necessary amount and mass out a sample of the metal salt.
4. Pour the samples of both metal salt and amino acid into the
pre-labeled Erlenmeyer flask and dissolve the mixture in 25
ml of water.
5. Obtain your pH checker and turn it on. Insert it into the amino
acid solution and allow it to stabilize. Once it has stabilized,
add enough 1 M HCl to it to adjust the pH of the solution to a
pH of about 1 to 1.5 (same value as in the previous
experiment). Record that pH and the volume of HCl added.
6. Obtain and clean a 50 ml buret. Condition the buret with a
standardized sodium hydroxide solution of concentration
approximately 0.1 M. Place the buret in a buret holder on a
ring stand and fill with the sodium hydroxide solution. Make
sure that there are no bubbles neither in the buret nor in its tip.
Record the initial volume reading in your notebook showing
two decimal places.
7. Add stepwise 0.5 ml of the NaOH solution to the studied
solution while stirring, and record pH after each addition (in
total 50 ml of NaOH should be added). Arrange your results
as a table:
V(NaOH), 0 0.5 1.0 1.5 2.0 … … … … …
ml
pH
8. Using the results of the titration, set up the titration curve (pH
vs. V(NaOH)) and add the titration curve obtained from the
previous experiment (see the figure above).
189
log***
logK2***
_
n ***
logK1***
log[L-]**
C(NaOH), mol/l*
C(M), mol/l*
C(HL), mol/l*
pH
V(NaOH), ml
No of the experiment
9. Basing on the mutual disposition of the two curves, conclude
on the ionic form of the ligand in the complex (neutral or
deprotonated).
10. Use the Hyperquad computer program to calculate stepwise
stability constants K1 and K2, and the overall formation
constant . You may also use the algorithm presented in the
table below:
1 2 3 4 5 6
7
8
9
10
1
2
...
...
* the dilution should be taken into consideration:
V (taken)
;
С  С (initial )
V (total)
** log[L-] = (pH – pK(COOH) + log(C(HL) – C(NaOH));
for pK(COOH) see the previous experiment;
*** see the theoretical part.
11. Calculate the mean K1, K2 and  values and mean deviations
using mathematical statistics methods.
190
EXPERIMENT 7
Determination of protonation constants of
nucleic bases by UV-spectrophotometry
Goals. Application of physico chemical methods (UVspectrophotometry) for determination of protonation constants of
organic species.
Theoretical part.
General approach.
Spectrophotometry is one of the most important practices
carried out in laboratories. Spectrophotometry is the study of the
reflection or transmission properties of a substance as a function
of wavelength. It consists of the qualitative and quantitative
studies.
The energy in the UV and visible regions ( 200 – 700
nm) irradiated on the molecules can result in changes in the
electronic nature of the molecule i.e. changes between ground
state and excited states of electrons within the system. As a
result, UV-visible spectroscopy is also known as electronic
spectroscopy.
As it was shown previously (see Lecture 1), the electrons
in the molecules occupy so called molecular orbitals (MO). The
electron transitions occur between occupied and unoccupied
MOs. The energy of the electron transition (i.e. E between the
occupied and unoccupied orbitals) is in accordance to the
absorbed wavelength:
191
R here is a Rydberg Constant (1.0974x107 m-1);  is the
wavelength; n is equal to the energy level (initial and final).
The electronic transitions in organic compounds can be
determined by ultraviolet-visible spectroscopy, provided that
transitions in the ultraviolet (UV) or visible range of the
electromagnetic spectrum exist for this compounds. The
longwawe absorption refers to the electron transition from the
higest occupied molecular orbital (HOMO) to the lowest
unoccupied molecular orbital (LUMO).
The change of the electronic structures of the molecules
because their tautomeric transformations, ionization (protonation
or deprotonation), complexation, ets. leads to the changes in
energies of HOMO and LUMO and, as a result, shifting of the
transition bands in the spectra.
Quantatively, the absorbance of light by a substance is
described by the Beer–Lambert–Bouguer law. It states that
there is a logarithmic dependence between the transmission (or
transmissivity), T, of light through a substance and the product of
theabsorption coefficient of the substance, α, and the distance the
light travels through the material (i.e., the path length), ℓ. The
absorption coefficient can, in turn, be written as a product of
either a molar absorptivity (extinction coefficient) of the
absorber, ε, and the molar concentration С of absorbing species
in the material:
,
where 0 and are the intensity (or power) of the incident light
and the transmitted light.
The transmission (or transmissivity) is expressed in terms
of an absorbance which, for liquids, is defined as
192
or
A  Cl .
Spectrophotometric titration.
Spectrophotometric titration is an analytical method in
which the radiant-energy absorption of a solution is measured
spectrophotometrically after each increment of titrant is added.
The studies of equilibrium processes are based on the difference
in absorption spectra of two forms of a substance (for example,
its neutral and ionized form). The existence of the equilibrium is
proved by the presence of one or several isobestic points:
Isosbestic point is a specific wavelength at which two
chemical species have the same molar absorptivity (ε) or, more
generally, are linearly related. The requirement for an isosbestic
193
point to occur is that the two species involved are related linearly
by stoichiometry, such that the absorbance is invariant for one
particular wavelength. The presence of an isosbestic point
typically does indicate that only two species that vary in
concentration contribute to the absorption around the isosbestic
point.
In case if an acid-base equilibrium of a weak acid or a
weak base is studied, it is evident that in acidic media a
protonated base or a neutral form of an acid are present in the
solution. The alkaline media are related to the neutral form of a
base and deprotonated form of an acid.
Thus, while titration of acidified solutions of a weak base
or a weak acid, the following processes takes place:
HA  H+ + AHB+  H+ + B
The equilibria constants of the processes can be expressed
as
Ka 
[ H  ][ A  ]
;
[ HA]
Kb 
[ H  ][ B]
;
[ HB  ]
In general, if  is considered as the degree of dissociation,
K
[H  ]
.
1
The  value at the each titration moment can be
calculated from the absorbance at a given pH. If the absorbance
in the acidified solution relates to the non-dissociated particles
(=0), and the absorbance in the alkaline solutions relates to their
full dissociation (=1), so
A  Aacidic
A  Aacidic
;
;

K  [H  ]
Abasic  Aacidic
Abasic  A
194
pK  pH  log
Abasic  A
A  Aacidic
Experimental part.
1. Prepare the necessary reagents and equipment:
Chemicals
Equipment
1 M HCl
Weighing boats
0.1 N NaOH
Spatula
Ethanol
Electronic pan balance
Nucleic base (assigned to each Macro (25 ml) and
student)
micropipets
pH 4 and 7 buffers
50 ml beaker
1-calibrated pH Checker
50 ml measuring flask
Magnetic rotator
UV-VIS spectrophotometer
2. Mass out a sample of your nucleic base and prepare 50 ml
of a 10-3 M solution in ethanol.
3. Record the UV-Vis spectrum of the sample according to the
common procedure (see the Instructions for the available
spectrophotometer). Dilute the solution if nessary to obtain
a good spectrum. Write down the necessary concentration.
4. Take 25 ml of the prepared solution in a beaker and
measure its pH. Add HCl dropwise until pH 1 is adjusted.
5. Adjust
your
spectrophotometer
program
for
spectrophotometric titration (see the Instructions).
6. Record the spectrum of the acidified solution. (Return the
solution back to the beaker after each spectrum
recordance).
195
7. Add 1 drop of the NaOH solution, stir it carefully with the
magnetic rotator, and measure pH (all measured pHs should
be added to the table (see below)). Record the spectrum.
8. Repeat the procedure 7 until pH 12 (or until there are no
more changes in the spectrum).
9. Save and print the results of the spectrophotometric
titration.
10. For calculations, select one or several analytical
wavelengths (they require to the maximal changes in
absorption within titration). As an example, see the Figure
above.
11. Measure absorbances at each titration point and arrange
the following table:
No of
pH A(1)
pK A(2) pK
…
experiment
1
2
…
12. Calculate pK value for all the intermediate points
(consider Ainitial as Aacidic and Afinal as Abasic.
13. Calculate the mean K1, K2 and  values and mean
deviations using mathematical statistics methods.
14. Search the corresponding pK value in the internet
databases and calculate the error of your experiment.
196
EXPERIMENT 8
Determination of stability constants of
complexes of transitional metals with the
nuclear bases by UV-spectrophotometry
Goals. Application of physico chemical methods (UVspectrophotometry) for determination of stability constants of
complexes of metals with nucleic bases.
Theoretical part.
In the spectrophotometric method, the molar absorptivity
of the complex is an additional variable to be determined. As
well, it is necessary to determine the stoichiometry of the
complex before calculations can be performed.
In the mole-ratio method a series of solutions is prepared
in which the concentration of one reactant (usually the ligand) is
held constant while the other reactant (usually the metal ion) is
varied.
The titration of the ligand solution by the solution of a
metal salt is performed, and the UV-Vis spectra are recorded after
each addition. The absorbance at the analytical wavelength is
then plotted versus the concentration of the metal in the solution:
197
If only one complex of high stability is formed, the graph
consists of two linear intersecting parts. The ratio of the
concentration of ligand at the intersection point to the (fixed)
concentration of metal ion gives the stoichiometry of the
complex.
If the stability constant of the complex is high (curve A),
there will be no appreciable dissociation of the complex at or near
the stoichiometric point. If the complex is moderately stable
(curve B), the plot will consist of two straight-line portions with a
central curved portion. Extrapolation of the two straight-line
portions yields the intersection point. If a complex of low
stability is formed, a large excess of ligand will have to be used to
drive the reaction to completion, and there will be no detectable
break in the curve from which to obtain the stoichiometry (curves
C and D).
198
Once the stoichiometry of the complex has been
established, the formation constant can be calculated provided the
data yields a curve showing some dissociation in the
neighborhood of the stoichiometric point (curve B). Briefly, for
any data point in the region of curvature, complex formation did
not proceed to completion, as evidenced from the difference
between the measured curve (B) and the “theoretical” one (A).
Here there is obviously an equilibrium between metal ion, ligand
and complex:
nMx+ + mL  MnLm.
The equilibrium constant of this process represents the
formation constant :
K  
 M n Lm ]
.
[ M x  ] n [ L] n
The equilibria concentrations of non-complexed particles
[Mx+] = CM – n[MnLm];
[L] = CL – m[MnLm]
(CM and CL are total concentrations of the metal ion and the
ligand in the equivalent point with respect to the mutual dilution
of solutions while mixing).
Considering a full transformation of the reactants
(“theoretical” curve A),
A(complex) = (complex) ℓ [MnLm]0
where
[MnLm]0 = 1/n CM.
(“theoretical” concentration)
So, the value of (complex) is calculated from the curve A.
Under the real conditions (experimental curve B), the
concentration of the complex
[MnLm] < [MnLm]0 .
From the experimental curve B which describes the
equilibrium process of complex formation, the real concentration
of the complex [MnLm] in the solution may be calculated:
199
A(equilibrium) = (complex) ℓ [MnLm]
(the (complex) value is already determined from the “theoretical”
curve A).
Now equilibria concentrations of the metal ion and the
ligand can be found as:
[Mx+] = CM – n[MnLm];
[L] = CL – m[MnLm].
And finally, knowing the composition of the complex and
equilibria concentrations of all the particles in the solution, the
formation constant of the complex can be easily calculated:

M n Lm ]
.
[ M x  ]n [ L ]n
Experimental part.
1. Prepare the necessary reagents and equipment:
Chemicals
Equipment
Ethanol
Weighing boats
Nucleic base (assigned to
Spatula
each student)
Metal salt (assigned to each Electronic pan balance
student)
Macro (25 ml) and micropipets
50 ml beaker
Two 50 ml measuring flasks
Magnetic rotator
UV-Vis spectrophotometer
2. Mass out a sample of your nucleic base and prepare 50 ml of
a solution of a nesessary concentration (see the previous
experiment).
200
3. Mass out a sample of the metal salt and prepare 50 ml of a
solution which is 100 times more concentrated than the
solution of a nucleic base.
4. Take 25 ml of the prepared solution of the nicleic base in a
beaker and record the UV-Vis spectrum according to the
common procedure (see the Instructions for the available
spectrophotometer).
5. Add stepwise 0.05 ml of the solution of a metal salt and
record the spectra after each addition (in total, you should
add 1 ml of the solution of a metal salt to 25 ml of the ligand
solution).
6. Select one or several analytical wavelengths (see the
previous experiment) and measure absorbance of each curve
in the titration.
7. Calculate the concentration of a metal salt at each point
(consider the dilution of the solution of the metal salt after
addition to the ligand).
8. Plot the absorbance at the analytical wavelength versus the
concentration of the metal in the solution, extrapolate the two
straight-line portions and determine the intersection point.
9. Knowing concentrations of metal and ligand, find the
composition of the complex in the solution.
10. Follow the procedure presented in the theoretical part and
calculate the stability constant of the complex compound.
201
EXPERIMENT 9
Determination of coordination sites of O- and
N-donating ligands by IR-spectroscopy
Goals. Application of physico chemical methods (IRspectroskopy) for determination of active sites of organic
compounds in the process of complex formation.
Theoretical part.
Infrared radiation is that part of the electromagnetic
spectrum between the visible and microwave regions. Infrared
radiation is absorbed by organic molecules and converted into
energy of molecular vibration, either stretching or bending.
Different types of bonds, and thus different functional groups,
absorb infrared radiation of different wavelengths. A IR spectrum
is a plot of wavenumber (X-axis) vs percent transmittance (Yaxis). (Note: wavelength can be used instead of wavenumber and
absorbance instead of percent transmittance).
A molecule or a functional group can vibrate in many
ways, and each way is called a vibrational mode. For molecules
with N atoms in them, linear molecules have 3N – 5 degrees of
vibrational modes, whereas nonlinear molecules have 3N – 6
degrees of vibrational modes. These are symmetric (a) and
antisymmetric stretching (b) or bending (c):
202
Complex IR spectra are interpreted by extrapolating from
such studies of simple molecules, since most functional groups
give rise to bands in particular regions of the spectrum. The
region from 4000-1300 cm-1 is particularly useful for identifying
the presence of specific functional groups: 3600 –2700 cm-1 for
X-H stretchings (C-H; O-H; N-H); 2700 – 1900 cm-1 for X≡Y
stretchings (C≡N; C≡C); 1900 – 1500 cm-1 for X=Y stretchings
(C=O; C=N; N=N; C=C); and 1500 – 500 cm-1 for X-Y
stretchings and X-H bendings.
Characteristic IR Absorption Frequencies of Organic Functional
Groups
Characteristic
Functional Type of
Absorptions
Intensity
Group
Vibration
(cm-1)
Hydroxyl
O-H
(stretch,
H-bonded)
3200-3600
strong, broad
203
O-H
(stretch,
free)
3500-3700
strong, sharp
C-O
(stretch)
1050-1150
strong
Amine
N-H
stretch
3300-3500
medium (primary
amines have two
bands; secondary have
one band, often very
weak)
C-N
stretch
1080-1360
medium-weak
N-H
bending
1600
medium
C-H
stretch
3000-3100
medium
C=C
stretch
1400-1600
medium-weak,
multiple bands
Aromatic
IR Absorption Frequencies of Functional Groups Containing a
Carbonyl (C=O)
Functional
Type of
Characteristic
Intensity
Group
Vibration Absorptions (cm-1)
Carbonyl
C=O
stretch
1670-1820
strong
(conjugation and coordination moves absorptions to lower wave
numbers)
204
Acid
C=O
stretch
1700-1725
strong
O-H
stretch
2500-3300
strong, very
broad
C-O
stretch
1210-1320
strong
C=O
stretch
1640-1690
strong
N-H
stretch
3100-3500
unsubstituted
have two bands
N-H
bending
1550-1640
C=O
stretch
1735-1750
strong
C-O
stretch
1000-1300
two bands or
more
Ketone
acyclic
stretch
1705-1725
strong
cyclic
stretch
3-membered –
1850
4-membered –
1780
5-membered –
1745
6-membered –
1715
7-membered –
1705
strong
,unsaturated
stretch
1665-1685
strong
aryl ketone
stretch
1680-1700
strong
Amide
Ester
205
Experimental part.
1. Prepare the necessary reagents and equipment:
Chemicals
Equipment
Ethanol
Weighing boats
Amino acid (assigned to each Spatula
student)
Metal acetate (assigned to each Electronic pan balance
student)
NaOH (diluted)
100 ml beaker
Nuyol
Heater
KBr or NaCl disks
Magnetic rotator
IR spectrophotometer
2. Peform the synthesis of the metal complex of one of the
amino acids according to the following procedure. Metal(II)
acetate (0.01 mole) is dissolved in 25 cm3 of hot water. 25
cm3 of hot ethanol is added and the solution kept hot. Amino
acid (0.02 moles) is dissolved in 25 cm3 of hot water. The
solutions are mixed while hot (~70C) and the solution then
cooled on ice. In the case if no precipitation is observed, the
pH of the mixture is adjusted to 7 – 8 with the help of the
diluted NaOH solution. After 2 hours, the formed precipitate
if filtered of (or centrifugated), washed with a small amount
of ethanol and dried in a dessicator over P2O5.
3. Prepare the dried sample of your complex as a suspention in
Nuyol between two KBr or NaCl disks and record an IR
spectrum according to a general procedure (consult the
Instructions for the IR spectrophotometer).
4. Using the procedure 3, record the IR spectrum of the noncoordinated amino acid.
206
5. Examine the IR spectrum of the amino acid. Using the tables
presented above, find the absorption bands which relate to
vibration modes of the amino- and carboxylic groups (how
many bands do you expect to describe and which vibration
modes they represent?).
6. Compare the spectra of the complex and the non-coordinated
amino acid and make your conclusions about the shifting of
the absorbtion bands while coordination.
207
208
Instructions for Research topic:
Presentation, Questions and Term Paper
During the first week of class, each student enrolled in the
course will choose a research topic out of the list included above
or propose his/her own topic related to the bioinorganic chemistry
for a class handout and presentation, as well as for a term paper.
(Note! Your research topic should be approved by the instructor).
Because the purpose is to have you learn something new, please
choose a topic you have not studied in a previous course.
The results of your researches should be arranged as a
Term Paper and Presentation.
The volume of the Term Paper should not exceed 10 –
15 pages. It should consist of the background information,
research problems, discussion, and conclusions. Include at least
one figure or table containing experimental data from your
research topic, and explain the interpretation of the data in the
figure or table. The list of citations should contain not less than 5
references: books or journal papers. avoid citations to web sites
unless you cannot find the information in a refereed journal
article (and in such a case make sure it is a trusted web site such
as one of those suggested by TEXT).
The class presentation should be a 10-minute “chalk
talk” accompanied by slides or a poster.The presentation and
paper may both start from the same outline, but because of the
brief time allocated to your presentation, you will need to
condense the background information and omit other recent
results in your presentation:
a) What is the biological function and/or medical relevance
of the system? In what species is this system found, and
209
from which species is the system best characterized? If
this a protein, is it part of a family of related proteins,
and if so, what are some of the names of the similar
proteins?
b) What is the metal involved?
c) What is the coordination environment of the metal?
d) What is known and generally accepted about the
chemical role of the metal and its coordination
environment in carrying out the function of the system?
Pay attention to the experimental results presented in your
Term Paper. What is learned about the system from the discussed
experiment?
In the conclusion, try to summarize what are some still
unanswered questions that are motivating further research on the
system that is the subject of your research topic.
Prepare two potential questions to ask your colleagues
during the discussion of your research topic, and be ready not
only to ask but to answer them.
210
Exam Questions
1. Draw the electronic configurations of the given atom and ion
and compare the relative stabilities of species. Indicate the
number of unpaired electrons.
2. Draw the electronic configurations of a given element in the
ground and possible excited states and predict its possible
valence states.
3. Using the VB method, describe the structures of the given
molecule and indicate the type of hybridization of the atoms.
4. Use the MO energy level diagrams to determine bond orders
and multiplicities of the ground states of the fgiven molecules
and ions and compare their relative stabilities.
5. Draw the MO energy level diagrams for the squaric (or
tetrahedric, or hexahedtic) complexes of one of the
transitional metal cation. Show the distribution of electrons in
the field of a strong (or weak) ligand.
6. Comparing the relative stabilities of complex ions conclude
on the possibility of their mutual transformations.
7. Describe the biological role of coordination compounds of Fe
(or Zn, or Cu, or V, or Cr, or Mo, or Co) in biological
processes.
8. Chelate effect in stabilisation of complexes by multidentate
ligands.
9. Pearson’s concept of hard and soft interactions in
coordination chemistry.
10. Protein structure.
11. Active sites of proteins. Coordination modes of amino acids.
12. A number of Copper (or Iron, or Zinc, or Cobalt) containing
enzymes are known. Give the names of two of these and show
the coordination environment around the metal.
13. The structures of iron centres of iron-sulphur proteins.
14. What are important properties of iron which can account for
its extensive use in terrestrial biological reactions?
211
15. Aspects of storage (or transport, or biomineralization) of iron
in the organisms.
16. Structure and characteristics of dioxygen-carrier proteins.
17. Electron structures of iron in deoxygenated and oxygenated
forms of hemoglobin.
18. Other than hemoglobin oxygen-binding proteins.
19. Requirements for Effective Oxygen Carriers.
20. Cooperative and non-cooperative binding of dioxygen.
21. Wigner's spin selection rule.
22. Dioxygen reactions.
23. Roles of metalloenzymes in dioxygen reactions.
24. General approach to dinitrogen biofixation.
25. The role of Mo- (or V, or all iron) nitrogenases in the process
of the dinitrogen fixation.
26. The role of quantum chemical modeling in bioinorganic
chemistry.
27. Theoretical approach to determination of ionization constants
of amino acids by potentiometric titration.
28. Calculation of stability constants of metal complexes with
weak acids by potentiometric titration.
29. Theoretical approach to determination of ionization constants
(deprotonation or protonation) of weak organic acids and
bases by spectrophotometric titration.
30. Calculation of stability constants of metal complexes with
organic ligands by spectrophotometric titration.
212
List of citations
1.
Ivano Bertini, Harry B. Gray. Bioinorganic Chemistry, Mill
Valley, California, 1994.
2. Glossary of terms used in Bioinorganic Chemistry. Ed. M.
W. G. de Bolster. Pure &App. Chem., Vol. 69, No. 6, pp.
1251-1303, 1997.
3. Lorenz Kienle. Basics of Bioinorganic Chemistry. MaxPlanck-Institut für Festkörperforschung. Stuttgart.
4. S.J. Lippard, J.M. Berg, Principles of Bioinorganic
Chemistry, U.S.B., 1994.
5. Bertini, I.; Gray, H. B.; Stiefel, E. I.; Valentine, J. S., editors.
Biological Inorganic Chemistry. University Science Books,
2007.
6. Housecroft, C.E. and Sharpe, A.G. “Inorganic Chemistry,
Pearson / Prentice Hall”, 2005 (2nd Edition).
7. Roat-Malone, R.M. Bioinorganic Chemistry: A Short
Course. Wiley, 2002.
8. Kazuya Arashiba, Yoshihiro Miyake Yoshiaki Nishibayashi
A molybdenum complex bearing PNP-type pincer ligands
leads to the catalytic reduction of dinitrogen into ammonia
Nature Chemistry, Vol. 3, 2011, pp. 120–125.
9. Jameson G.B. and Ibers J.A. Biological and Synthetic
Dioxygen Carriers.
http://authors.library.caltech.edu/25052/5/BioinCh_chapter4.
pdf
10. Casiday R. and Frey R. Hemoglobin and the Heme Group:
Metal Complexes in the Blood for Oxygen Transport.
http://www.chemistry.wustl.edu/~edudev/LabTutorials/Hem
oglobin/MetalComplexinBlood.html
11. http://simple.wikipedia.org/wiki/
12. http://sandbian.wordpress.com/articles-papers/
213
13. http://www.adichemistry.com/inorganic/bioinorganic/bioinor
ganic-chemistry.html
14.
Billo, E.J. (1997). "22". Excel for chemists: a
comprehensive guide (2nd ed.). Wiley-VCH. ISBN 0-47118896-4. (Electronic)
http://www.ahut.edu.cn/yxsz/ahkl/Teaching/Excel%20for%20
Chemists/Ch22.pdf.
15.
A. Albert und E. P. Serjeant: Ionization Constants of
Acids and Bases. 1. Auflage. Methuen & Co., London, John
Wiley Sons, New York 1962. 179.
16.
Beck, M.T.; Nagypál, I. (1990). Chemistry of Complex
Equilibria. Horwood. ISBN 0-85312-143-5.Chapter 1.
17.
Rossotti, F.J.C.; Rossotti, H. (1961). The Determination of
Stability Constants. McGraw–Hill.
18.
Koji Nakanishi, Philippa H. Solomon. Infrared Absorption
Spectroscopy.
19.
Silverstein, R.M.; Bassler, G.C.; and Morrill, T.C.
Spectrometric Identification of Organic Compounds. 4th ed.
New York: John Wiley and Sons, 1981.
20.
http://facstaff.bloomu.edu/jmorgan/amino%20acid%20det
ermination%20Spr09.pdf
21.
http://orgchem.colorado.edu/Spectroscopy/irtutor/tutorial.
html
22.
http://www.chem.ucla.edu/~webspectra/irintro.html
23.
http://en.wikipedia.org/
24.
http://www2.ups.edu/faculty/hanson/Spectroscopy/IR/IRfr
equencies.html
214