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Transcript
NAME: IDU DOREEN
MATRIC NO: 14/SCI03/011
COURSE: BIOINORGANIC CHEMISTRY
DATE: 14-11-2016
Metalloenzymes remain an active area of research due in part to successes in targeting these
enzymes for drug development. An estimated 10% of approved drugs target metalloenzymes,
including angiotensin-converting enzyme, carbonic anhydrase, and matrix metalloprotesases.
Since inhibitors of metalloenzymes often contain a metal-binding group that targets the
catalytic metal ion, the interaction between the metal-binding group and catalytic metal ion
will significantly impact inhibitor affinity for the target enzyme. Therefore, the development
of therapeutically effective inhibitors requires proper identification of the biologically
relevant metalloenzyme cofactor(s). The roles of metal ions in enzyme active sites (aside
from structure maintenance) include electron transfer, oxygen atom transfer, formation of
coordinated hydroxide, electrophilic catalysis, as well as substrate binding. Metalloenzymes
catalyze numerous reactions of physiological importance, including mitochondrial O2
reduction, peptide bond cleavage, hydrocarbon hydroxylation, destruction of O2− and H2O2,
and hydration of CO2.
Metalloenzymes play key roles in many processes central to human physiology, including the
biosynthesis of DNA and certain amino acids, steroid metabolism, destruction of superoxide
and hydrogen peroxide, biosynthesis of leukotrienes and prostaglandins, carbon dioxide
hydration, neurotransmitter metabolism, digestion, collagen biosynthesis, and, of course,
respiration. The latter could be viewed as a process of global bioenergetic importance, one
that complements photosynthesis – the dioxygen that is evolved (via the Mn4 oxygen-
evolving complex of Photosystem II) by photosynthetic organisms is consumed by aerobic
microbes and animals (Fe, Cu cytochrome oxidases catalyze the reduction of O2 to H2O).
It has become widely appreciated that metal ions play important catalytic roles that frequently
cannot be matched by protein side chains or organic prosthetic groups. The most significant
observation of this type concerns the microbial fixation of molecular nitrogen, a very inert
molecule:
N2 + 6H+ + 6e− −−−→ 2NH3
This reaction is catalyzed by nitrogenase, whose active site consists of a dissociable Mo- and
Fe-containing cofactor. The catalysis of such multielectron redox reactions is frequently
carried out by metalloenzymes containing multimetal centers (i.e. metal clusters).
Metal Ion Bioavailability
Only a small number of the metallic elements appear to be utilized in biology. Of these, Na+,
K+, Ca2+, and Mg2 are considered macro minerals, or bulk elements; high concentrations of
these ions are needed for osmotic homeostasis, neuromuscular transmission, and bio
mineralization (e.g. bone formation). Other essential metals are present in trace quantities in
humans and most other organisms. Even the most prominent biologically active transition
metals (iron, zinc, and copper) are trace elements. The selection of metal ions for
incorporation into metalloenzymes is strongly influenced by bioavailability – a given element
must be abundant in the environment and must be present in an extractable form. A striking
exception to this generalization involves the nearly universal requirement for iron. Organisms
have evolved selective uptake mechanisms for this element, the most abundant transition
metal in the earth’s crust, which forms insoluble ferric hydroxides in the presence of O2. The
incorporation of metal ions into metalloenzymes is also influenced by other, chemically
oriented, parameters such as ionic radius, charge, preferred coordination geometry, ligand
substitution and redox kinetics, aqueous solution chemistry, and thermodynamic stability.
Metal/Ligand Substitution
Metalloenzymes frequently contain metal cofactors that are spectroscopically useless. For
example, zinc enzymes are diamagnetic and colourless – electron spin resonance or electronic
absorption spectroscopy are therefore uninformative. However, information about the
environment of the metal-binding site can be obtained by replacing the Zn2+ ion with a
paramagnetic, chromophoric metal-ion probe. Co2+ readily replaces Zn2+ – they prefer
similar coordination environments, have comparable ionic radii, and display similar reactivity
patterns. The resultant cobalt-substituted enzyme can be characterized with regard to
intermediates formed during enzymatic turnover. Furthermore, the use of metal derivatives
can also indicate whether the metal binding site is rigid or flexible. Obtaining the structure of
a metalloenzyme is only the first step in developing an understanding of its physiological
function. The production of recombinant derivatives affords opportunities to address
additional issues, such as endogenous ligand exchange at the active-site metal(s) and
participation of nearby amino acid residues in substrate binding/release, proton transfer,
and/or electron transfer. Fast kinetic techniques (stopped-flow spectrophotometry,
freezequench trapping of intermediates, flash photolysis, temperature-jump relaxation) are
typically used to map out the individual steps that characterize the turnover of a
metalloenzyme. In many cases, individual steps are too fast to accurately measure. One
commonly used application of such techniques is the study of absorption spectra of transient
enzyme intermediates generated by photolyzing carbon monoxide adducts of reduced heme
enzymes in the presence of O2. What tends to be overlooked in interpretations of structure
and reactivity data is theimportance of protein fluctuations (particularly in substrate binding
and product release) – metalloenzymes are much more than metal ions in disguise.
Functional Roles of Metal Cofactors
There are two principal types of metal dependent enzyme. Metalloenzymes contain at least
one tightly bound metal ion, at the active site, that is required for activity. Metal-activated
enzymes, on the other hand, generally lose catalytic activity during purification because their
affinity for the required metal is rather low. Mg2+-, K+-, and (most) Ca2+-dependent
enzymes are metal-activated and will not be discussed further. Na+-dependency has yet to be
unequivocally demonstrated for an enzyme. Examples of metalloenzymes are known for each
class of enzyme designated by the International Union of Biochemistry and are as follows:
oxidoreductases, transferases, hydrolases, lyases, isomerases, and ligases. The following
properties of metal ions make them well suited as catalysts for these types of reactions:
1. Metal ions bind at least three (usually four or more) ligands, thereby promoting the
organization of protein structure.
2. With the notable exception of zinc, all of the other known transition metal cofactors can
potentially exist in two or more oxidation states; metalloenzyme catalysis of oxidationreduction (redox) reactions is thus quite common.
3. Metal-ion cofactors are electrophilic and can serve as effective Lewis acids for binding and
activating substrates. Two types of metal-ion reactivity are of fundamental importance in
considering the wide range of activities displayed by metalloenzymes: changes in oxidation
state and changes in bound ligands. Numerous examples of electron-transfer metalloproteins
(blue copper proteins, iron–sulfur proteins, cytochromes) are known; the metal coordination
spheres of these electron carriers do not change during electron transfer. Metal ions
possessing static coordination spheres could alternatively play purely structural roles, as do
Ca2+ in calmodulin, horseradish peroxidase, or stromelysin. Certain metals (e.g. Zn2+ in
carboxypeptidase A) cannot undergo redox reactions within the constraints imposed by
nature; instead, acid–base catalysis is the raison d’ˆetre for the metal in the enzyme. Lastly,
both the metal-ion oxidation state and coordination sphere could change during enzymatic
turnover. Regardless, it is clear that metal ions that directly participate in enzymatic catalysis
must possess dynamic coordination environments.
References
1. Morrison C (2009) Financings of the Fortnight: Summer Madness, The In Vivo Blog,
Elsevier, Inc.
2. Bertini, I., Gray, H.B., Lippard, S.J., Valentine, J.S., (Eds.) (1994) Bioinorganic
Chemistry, University Science Press, Mill Valley, CA.
3. Messerschmidt, A., Bode, M., Cygler, W. (Eds.) (2004) Handbook of Metalloproteins,
Encyclopedia of Molecular Cell Biology and Molecular Medicine, 2nd Edition.
Volume 8
4. Edited by Robert A. Meyers.
-VCH Verlag GmbH & Co.
KGaA, Weinheim ISBN: 3-527-30550-5Vol. 3, Wiley, New York