Download 2. CYCLIC AMINOACIDS 2.1. Aromatic

Medical Biochemistry
Molecular Principles of Structural Organization of Cells
4. PROTEINS
PROTEINS
The major cell components of any living organism (25% of wet
weight and 45-50% of dry weight)
Play important roles in all biological processes
Elementary composition: C 51-55%, O 21-23%, N 15-18%, H 6-7%, S 0.3-2.5%
Structure - they are
– high-molecular (the mass of single-chain protein is 10-50 kilodaltons
(350 dal-1000 kdal); multichain protein complexes >200 kdal.
– N containing organic compounds (16% of dry weight),
– with complex structural organization,
– constructed from 20 different aminoacids,
– linked in chains by peptide bonds.
Depending on the chain length peptides are classified in
– Oligopeptides = 2-10 aa
– Polypeptides = 10-40 aa
– Proteins = >40 aa
NATURE OF PROTEINS
Functions:
– Enzymatic catalysis
– Transport and storage of small molecules and ions
– Structural (cytoskeleton), providing strength and structure to cells,
forming components for intracellular and extracellular movements
– Immune defense system (antibodies)
– Hormonal regulation (hormones and receptors)
– Control of genetic expression – activators, repressors
Show specificity of biological function, as a consequence of
the uniqueness of three-dimensional structure
AMINOACIDS
– STRUCTURAL MONOMERS OF PROTEINS
Aminoacids contain at least 1 –NH2 group and 1 –COOH group.
L-aminoacids are classified in α-, β-, γ- depending on the
position of C bearing –NH2 group with respect to –COOH.
There are >200 aa in different species, 60 in human, only 20 in
the structure of proteins.
Aa are classified in:
– proteogenic - in the structure of proteins
– nonproteogenic – not incorporated in proteins
Three classifications are adopted:
– Structural
– Electrochemical
– Biological (physiological)
All protein aa are L-aminoacids and α-aminoacids
AMINOACIDS – FUNDAMENTAL UNITS OF PROTEINS
STRUCTURAL CLASSIFICATION
1. ACYCLIC
1.1. Aliphatic unsubstituted
Glycine (Gly)
Alanine(Ala)
O
H2N
CH C
H
O
OH
H2N CH C OH
H2N CH C OH
Leucine (Leu)
O
H2N
CH3
1.2. Aliphatic substituted
1.2.1.Hydroxy aa
Serine (Ser)
O
(hydroxyamine a)
1.2.2.Thio- aa
(thiamin a)
Valine (Val)
Isoleucine (Ile)
O
CH C OH
H2 N
CH C
O
OH
H2 N
CH2
CH CH3
CH3
CH CH3
CH2
CH3
CH3
O
CH C
OH
CH2
CH OH
OH
CH3
Cysteine (Cys) Cystine (Cys2) Methionine (Met)
O
H2N CH
C OH
O
O
H2N
CH
C OH
H2N
CH C
CH2
CH2
CH2
SH
S
CH2
S
S
CH2
CH3
H2N
OH
CH CH3
Threonine (Thr)
H2N
CH C
CH
COOH
OH
1.2.3. Monoamino- Aspartic acid Glutamic acid Asparagine Glutamine
dicarboxylic
(Asp)
(Glu)
(Asn)
(Gln)
O
O
O
O
(carboxyamine a)
H2N
CH C
OH
H2N
CH C
OH
H2 N
CH C
OH
CH2
CH2
CH2
C
CH2
C
CH2
C
NH2
O
O
O
Lysine (Lys)
CH2
CH2
CH2
CH2
NH2
Arginine (Arg)
O
H2N
CH C OH
CH2
CH2
CH2
NH
C
NH
NH2
O
Hydroxylysine (Lys-OH)
O
O
H2N CH C OH
C
NH2
OH
1.2.5. Guanidine
amine
acids
CH C
CH2
OH
1.2.4. DiaminoMonocarboxylic
(diamine acids)
H2N
H2N
CH C OH
CH2
CH2
CH OH
CH2
NH2
Aminocitric acid
H2N CH COOH
OH
OH
C COOH
H2C COOH
2. CYCLIC AMINOACIDS
2.1. Aromatic aa
Phenylalanine Tyrosine
(Phe)
(Tyr)
O
H2N
CH C
O
OH
CH2
H2N
CH C
OH
CH2
OH
2.2. Heterocyclic aa
Histidine
Tryptophan
(His)
(Trp)
O
H2N
CH C OH
Proline Hydroxyproline
(Pro) (Pro-OH)
O
H2N
CH2
CH C
O
OH
C
OH
C OH
CH2
HN
N
NH
O
HN
HN
OH
Rare aminoacids: Aminocitric acid, Lys-OH, Pro-OH
AMINOACIDS - STRUCTURAL CLASSIFICATION
ACYCLIC AMINOACIDS
1.1. Aliphatic unsubstituted:
1.
•
•
Glycine/glycocol: excretion of benzoic acid as benzoylglycine, constituent of
glutathione, intermediate in the synthesis of creatine, hem, purine bases
Alanine, valine, leucine, isoleucine: nonpolar, hydrophobic bonds
1.2. Aliphatic substituted:
1.2.1. Hydroxyamine acids:
–
Serine: slightly acidic; role:
–
–
–
–
–
constituent of active sites of some enzymes,
binding site of olygosaccharides in glycoproteins
Phosphoserine in phosphoproteins (phosvitin, vitellin, casein, myosine), phosphorylated
enzymes,
in phosphatidylserine
Threonine: slightly acidic; role:
–
–
–
active site of enzymes
binding site of olygosaccharides in glycoproteins
phosphothreonine in phosphoproteins (casein, tropomyosin)
1.2.2. Thiamin acids:
Cysteine: slightly acidic, converted by oxidoreduction in cystine, forms
disulfide bonds between peptide chains; role: in the structure of
glutathione, metallothioneines, excretion of aromatic substances
Cystine: reduced to cysteine; in the structure of keratin, hair, insulin
Methionine: nonpolar, furnishes the 8 atoms of C in cysteine synthesis;
crystalline in the lens contains N-acetylmethionine
1.2.3. Carboxy acids:
Aspartic acid: enzymes active sites, urea cycle, synthesis of nitrogenous
bases
Glutamic acid: glutathione, folic acid, collagen; transamination,
glutaminogenesis
1.2.4. Diamine acids:
Lysine: cationic at pH 7; binds cofactors at the active site of enzymes
Lysine-OH: collagen, bonding site for olygosaccharides
1.2.5. Guanidinamine acids:
Arginine: basic, binds phosphate group; takes part in urea cycle,
biosynthesis of creatine
2. CYCLIC AMINOACIDS
2.1. Aromatic:
Phenylalanine: nonpolar, in the synthesis of tyrosine
Thyrosine: slightly acidic; enzyme bonding with substrate, synthesis of
tyroxine, catecholamines, melanins
2.2. Heterocyclic:
Histidine: active site of enzymes, binds metal ions, in the structure of
anserine and carnosine (dipeptides); generated histamine
Tryptophan: precursor of serotonin, crystalline in lens
Proline: role in folding the polypeptide chain
Proline-OH: collagen, elastin, acethylcolinesterase
AMINOACIDS – ELECTROCHEMICAL
CLASSIFICATION
Acidic – additional -COOH groups in the sidechain, polar:
– aspartic acid,
– glutamic acid,
– aminocitric acid
Basic - cary additional basic groups: amino, guanidine, imidazole), polar:
– lysine,
– arginine,
– histidine
Neutral - nonpolar, hydrophobic acids
AMINOACIDS – BIOLOGICAL CLASSIFICATION
Essential (8) - cannot be synthesized in the organism
– Val, Leu, Ile, Thr, Lys, Met, Phe, Trp
Half-essential (3) - can be synthesized not in sufficient amounts
– Arg, Tyr, His
Nonessential - can be synthesized by the organism
NONPROTEOGENIC AMINOACIDS
ornithine and citrulline – intermediates in urea cycle, synthesis of arginine
γ-aminobutyric acid (GABA) – free in the brain, lungs, heart;
neurotransmitter
β-alanine – in the strucutre of vitamin B3, CoA-SH, carnosine and anserine;
product of pyrimidines catabolism
dihydroxyphenylalanine (DOPA) – intermediate in the synthesis of
hormones of adrenal medulla
O
CH2 NH C NH2
H2N
CH2 NH2
CH2
CH2
CH2
CH2
CH2
CH2
CH2
CH2 NH2
CH NH2
CH
CH2
CH2
COOH
COOH
COOH
COOH
ornithine
NH2
citrulline
NH2
γ-aminobutyric acid
GABA
CH COOH
β-alanine
OH
OH
DOPA
AMINOACIDS – PHYSICAL AND CHEMICAL
PROPERTIES
Acid-base properties
Aa have amphoteric properties (have both acidic and basic groups)
Monoamino-monocarboxylic aa exist in aqueous solutions as zwitterions
(dipolar molecules): carboxyl is dissociated and negatively charged,
amino is protonated and positively charged; they are electrically neutral.
1.
–
–
At low pH COO- accepts H+ and becomes uncharged; the molecule is positive
At high pH the NH3+ loses H+ and becomes uncharged; the molecule is negative
The aminoacids having side chains that contain dissociating groups:
–
–
–
Aspartic acid, glutamic acid are acidic
Lysine, arginine, histidine are basic
Cysteine, tyrosine have a negative charge on the sidechain when dissociated
The state in which the net charge on the aa is 0 = isoelectric point (pHi) a
very accurate indicator of acid-base properties:
–
–
–
–
for nonpolar aa – close to neutral (5.5 for Phe, 6.3 for Pro)
for acidic aa – low values (3.2 for Glu)
weak acidic – for Cys, Cys-S-S-Cys (5)
for the rest, especially Lys, Arg, His – higher than 7
2. All proteogenic aa except glycine have an asymmetric C*, exibiting optical
activity. They exist as stereoisomers or enantiomers (L- or D-)
R
R
l
l
H2N – C* – H
H – C* – NH2
l
l
COOH
COOH
L-aminoacid
D-aminoacid
All the native aa are levorotatory as they rotate to the left the plane-polarized
light; they belong to L- series
D-aminoacids exist in bacterial products (cell walls), peptide antibiotics, but
not incorporated in proteins via ribosomal synthesis
STRUCTURE AND
LEVELS OF STRUCTURAL ORGANIZATION OF PROTEINS
PRIMARY STRUCTURE
The simplest level of structural organization = a linear polypeptide chain that is
composed of aminoacids radicals linked through covalent peptide bonds formed
between the α-amino group of one aa and the α-carboxyl group of the next aa.
R1
R2
R1
R2
l
l
l
l
H2N-CH-COOH + H2N-CH-COOH → H2N-CH-CO-HN-CH-COOH
-H2O
peptide bond
aminoacid1
aminoacid2
dipeptid
Specific characters of the peptide bond:
– coplanarity (all the atoms – CO-NH- in a single plan)
– two resonance forms (keto- and enol)
– trans position of the substituent to C-N bond
– ability to form H bonds
O
R2
ll
l
- CH - C – N – CH l
l
R1
H
Nomenclature for peptides:
– 2 aa (aa residues or radicals) = dipeptide;
– 3 aa = tripeptide; and so on
examples:
carnosine
= β-alanyl-histidine
anserine
= β-alanyl-N-methyl-histidine
glutathione = γ-glutamyl-cysteinyl-glycine
– synthesized in the erythrocytes, liver, intestinal mucosa, brain;
– a systemic protectant against oxidative stress, detoxification
from peroxides, cofactor for antioxidative GPx enzyme,
transmembrane transport, receptor action, antitoxic
– takes part in redox processes, coenzyme that donates H,
activator of SH-dependent enzymes,
– Polypeptides > 10 aa residues
– Proteins > 40 aa residues
All peptides or proteins contain:
R1
R2
R3
R4
R5
R6
l
l
l
l
l
l
H2N-CH-CO-HN-CH-CO-HN-CH-CO-HN-CH-CO-HN-CH-CO-HN-CH-COOH
N-terminal aa = free -NH2
C-terminal aa = free –
COOH
(written to the left)
(written to the right)
Aa are named consecutively beginning with the N-terminal aa,
bearing the suffix –yl, except the C-terminal aa that has its
name ended in –ine
(e.g. valinyl-leucinyl-alanine)
STRUCTURE AND
LEVELS OF STRUCTURAL ORGANIZATION OF PROTEINS
SECONDARY STRUCTURE
Refers to the way the peptide is folded into an ordered
structure owing to hydrogen bonds between the peptide
groups of the same or juxtaposed polypeptide chain
Classified in:
 -helix
 -structure
SECONDARY STRUCTURE
1. helical structure (α-helix):
• helical configuration, right-handed
(clockwise turns)
• H bonds are formed between
peptide groups within the same
polypeptide chain, between the 1st
and 4th aminoacid radical; there are
3.6 aminoacid residues per turn
• regularity of turns along the helix
length
• equivalence of all aa residues
(irrespective the R structure)
• nonparticipation of R groups in H
bonding
Barker R: Organic Chemistry of Biological Compounds, Englewood Cliffs, NJ, Prentice Hall, 1971
SECONDARY STRUCTURE
2. pleated sheets (β-structure):
the chains lie side by side,
with the H bonds formed
between the
– -CO- group of one peptide
bond
– –NH- group of another
peptide bond in the
neighboring chain
the chains may run
– in the same direction
(parallel β-sheet) or
– in opposite direction
(antiparallel β-sheet)
Barker R: Organic Chemistry of Biological Compounds,
Englewood Cliffs, NJ, Prentice Hall, 1971
SECONDARY STRUCTURE
The α-helix can be reversibly converted to β-structure due to the
reorganization of the H bonds (e.g. keratin, the protein in hair)
The same protein has both types of structure:
paramyosin has 95-100% α-helix,
myoglobin, hemoglobin, have high percentage of α-helix;
keratin, collagen (skin, tendon) have β-structure
SECONDARY STRUCTURE
3.
Collagen triple helix
- Constituent of skin, bones, teeth, blood vessels, tendons, cartilage,
connective tissue, the most abundant protein in the human (30% of
total body mass)
- Contains 33% Gly, 21% Lys-OH or Pro-OH, almost absent Cys
- The tropocollagen structure (the repetitive unit) is formed of 3 protein
strands that wrap around each other forming a left-handed superhelix,
held together by hydrogen bonds formed by the –OH in the Lys-OH or
Pro-OH
- 10 different types: I in tendons and bones, II in hyaline cartilage, III
in connective tissue, IV in basement membranes, VI in placenta
STRUCTURE AND
LEVELS OF STRUCTURAL ORGANIZATION OF PROTEINS
TERTIARY STRUCTURE
Is referred to as a specific mode of spatial arrangement of the polypeptide
chain: globular (ellipsoidal shape) and fibrous species (elongated)
Due to the associations between segments of α-helix and β-structure,
representing a state of lowest energy and greatest stability
Bonds formed between the sidechain radicals of aminoacids stabilize the
structure:
– Strong bonds:
Covalent
– Disulfide (-S-S-)
– Isopeptide (peptide-like, -CO-NH-)
– Ester (-CO-O-)
– Weak bonds:
Polar bonds:
– Hydrogen bonds
– Ionic or electrostatic
Nonpolar bonds (van der Waals)
Barker R: Organic Chemistry of Biological Compounds,
Englewood Cliffs, NJ, Prentice Hall, 1971
TERTIARY STRUCTURE
Specific features:
– The conformation is determined by the properties of the sidechain
radicals and medium
– The molecule tends to adopt an energetically favorable configuration
corresponding to the minimum of free energy
The nonpolar R form an interior region with hydrophobic radicals
The polar, hydrophylic R extend outside, oriented to the water
molecules
There are regions formed as α-helix or β-structure and random coils
The tertiary structure determines the protein activity
STRUCTURE AND
LEVELS OF STRUCTURAL ORGANIZATION OF PROTEINS
QUATERNARY STRUCTURE
Represents the aggregation of 2 or more polypeptide chains
(protomers or subunits) with tertiary structure, organized into
a single functional protein molecule, named oligomer.
Configuration of their tertiary structure, globular or fibrous.
Contacts between the subunits are possible through the polar
groups in R, as the nonpolar aminoacids radicals are oriented
to the interior
Bonds:
– Weak:
ionic bonds
hydrogen bonds
– Covalent:
disulfide
Examples:
hemoglobin (Hb) the blood pigment is a tetramer, constituted
of 4 protomers: 2 identical α-chains, 2 identical β-chains. The
four protomers form 2 subunits (αβ). The association can be
represented:
2 α + 2 β → α2β2 → 2 αβ
allosteric enzymes – phosphorylase a is a dimer, formed of 2
identical subunits (that separately are inactive)
PHYSICAL AND CHEMICAL PROPERTIES OF
PROTEINS
1. Amphoteric – as they combine acidic and basic properties
due to the acid-base groups of the side-chain radicals of the
protein constituting aminoacids.
the majority of the polar groups are located on the surface of
globular proteins, providing the acid-base properties and the
charge of the protein molecules
– Acidic aminoacids (glutamic, aspartic, aminocitric) → acidic properties
– Basic aminoacids (lysine, arginine, histidine) → basic properties
Buffering properties – the proteins containing a large amount
of histidine radicals, because its side-chain exibit buffering
properties within a pH range close to the physiological pH, for
example hemoglobin (8% histidine)
2. Colloidal and osmotic properties: aqueous solution of proteins
are stable and equilibrated (do not precipitate), homogeneous.
Properties of colloidal solutions:
–
–
–
–
–
Characteristic optical properties
Low diffusion rate
Inability to pass across semipermeable membranes
High viscosity
Property of gelation
3. Hydration of proteins and factors affecting solubility
Proteins are hydrophilic
Factors affecting solubility
– The charge on protein molecules (the higher the number of polar
aminoacids – the greater the amount of water bound)
– Neutral salts in small concentrations enhance the solubility
– The medium pH values
– Temperature influences differently, depending on the specific protein
Salting-out is the selective precipitation of a protein by a
neutral salt solution, used for separation and purification of
proteins; after removing the salting-out agent, the protein
retains its native properties and functions unchanged.
Denaturation and renaturation: agents destroy the higher
levels of structural organization of protein molecules
(secondary, tertiary, quaternary) by the breakdown of bonds
that stabilize them and retention of the primary structure; as
the result, the protein loses its native physical-chemical and
biological properties = denaturation; the protein separates
from solution as a precipitate.
Factors producing denaturation:
–
–
Physical: temperature, pressure, mechanical action, ultrasonic, ionizing
irradiation
Chemical: acids, alkali, organic solvents, detergents, certain amides (urea,
guanidine)alkaloids, heavy metal salts (Hg, Cu, Ba, Zn, Cd)
Properties of denaturated proteins:
1.
2.
3.
4.
5.
An increased number of reactive and functional groups – the unfolding of
polypeptide chain
Reduced solubility, increased ability to precipitate
Alteration of configuration
Loss of biological activity
A facilitated cleavage by proteolytic enzymes
Denaturation is used to deproteinize a mixture of protein-containing materials.
Removing the proteins one can obtain a protein-free solution
Denaturation was thought to be irreversible; in certain conditions the protein
restores its biological activity = renaturation
CLASSIFICATION AND NOMENCLATURE OF
PROTEINS
Physical-chemical classification:
– Electrochemical properties:
Acidic (polyanionic proteins)
Basic (polycationic proteins)
Neutral
– Polar properties
Polar / hydrophylic
Nonpolar / hydrophobic
Amphipathic / amphyphylic
Functional classification – biological functions
Structural classification
– Simple/unconjugated/apoproteins = polypeptide chain
– Conjugated/proteids = polypeptide chain + nonprotein moiety
(glycoproteins, lipoproteins, phosphoproteins, nucleoproteins,
metalproteins, cofactor-proteins)
SIMPLE PROTEINS
1.
2.
3.
4.
5.
Histones
– form reversible complexes with DNA = chromatin; histone-like proteins
exist in ribosomes
– stabilize the spatial structure of DNA and chromosomes
– interrupt the genetic information transfer from DNA to RNA
Protamines
– the most low-molecular proteins, basic, bound to DNA in the chromatin
of spermatozoa
Prolamines
– plant proteins in grain gluten of cereals: gliadin of wheat, avenin from
oats, zein from corn
– nonpolar aminoacids and proline - insoluble in water, salt solution, acid
and bases
Glutelins
– plant proteins, high content of arginine, low content of proline insoluble in water, salt solution, ethanol; soluble in diluted alkali,
Scleroproteins
– bones, cartillage, ligaments, tendons, nails, hair
– fibrous protein soluble in special solvents
6. Albumins and globulins are heterogeneous groups of proteins contained in
the blood plasma, in cells and biological fluids, with highly diversified
functions.
Albumins:
Relatively small molecular mass (15,000-17,000 Daltons)
Possess a negative charge
Acidic properties (isoelectric point 4.7); high content of glutamic acid
Strongly hydrated – are precipitated only at high concentrations of waterabsorbing agents
High absorbtive capacity for both polar and nonpolar molecules (transport
agents)
Globulins:
Higher molecular mass (>100,000)
Insoluble in pure water, soluble in dilute salt solutions
Weakly acidic or neutral (isoelectric point 6-7.3)
Weakly hydrated – are precipitated in low-concentrated solutions of
ammonium sulfate
Some of them specifically bind various materials (specific transport agents)
others nonspecifically bind lipid-soluble materials
Can be separated by electrophoresis because they have different
mobility under an applied electric field.
Albumins are polyanionic proteins and move to the anode faster
than globulins
Globulins are divided into 3 major fractions α (α1, α2), β (β1, β2), γ
CONJUGATED PROTEINS
Heteromacromolecules = macromolecular complexes composed of
2 components of different chemical classes.
Conjugated proteins (protein-nonprotein complexes)
1.
Nucleoproteins = proteins + nucleic acids
1. DNA-protein complexes (DNA + histones/nonhistones) =
deoxyribonucleoproteins (DNP) in the chromosomes
2. RNA-protein = ribonucleoproteins (RNP), in the cell
2. Glycoproteins = proteins (80-90%) +
•
•
•
•
heteropolyglucide
Have higher thermal stability
Difficult to be digested by proteolytic enzymes (pepsin, trypsin)
Exist in the blood, cell membrane (with the carbohydrate residue always
located on the external surface), inside the cell
Biological functions:
Transport of hydrophobic materials and ions (ceruloplasmin, transferrin,
haptoglobin,transcortin)
Blood coagulation (prothrombin, fibrinogen)
Immunity (immunoglobulins)
Enzymes (cholinesterase)
Hormones (corticotropin, gonadotropins)
Specificity of intercellular contacts - on the membrane surface – act as
recognition and binding sites (receptors) for the substances to be taken
up by the cell
Blood-group specificity - on the surface of the erythrocytes, are antigens
that determine whether an individual has type A (N-acetyl
galactosamine), B (D-galactose), AB (both) or O (absence of both) blood
Mucus secreted by the epithelial cells lubricates and protects the tissues
lined by these cells
Lipoproteins = lipids (triglycerides, cholesterol, cholesterides, phospholipids)
3.
–
–
–
–
+ proteins (apolipoproteins)
The high content in lipid determines the higher molecular mass and lower
density
The apolipoprotein differ in structure and composition: A1, A2, A3, B, C1, C2,
C3, D, E
Micelles-like structure:
hydrophobic core of nonpolar lipids (triacylglycerides, cholesterol esters)
hydrophylic envelope of polar lipids (cholesterol, phospholipids) and
proteins
By ultracentrifugation (or electrophoresis) the lipoproteins are separated in:
Proteins
Chylomicrons,
2%
Very Low Density Lipoproteins(VLDL)/ pre-β-lipoproteins 5-10%
Intermediate Density Lipoproteins (IDL)
15-20%
Low Density Lipoproteins (LDL) / β-lipoproteins
20-25%
High Density Lipoproteins (HDL)/ α-lipoproteins
45%
–
Major lipid
TG
TG
TG, C, PL
C
PL, C
Function
transport exogeneous TG
transport TG livertissues
transport C livertissues
transport C tissues  liver
Functions:
Structural – biological membranes providing the physiological function of
cells, nerves and transport of materials
Plasmatic - transport of lipids supplied by the intestinal absorption and
their distribution among lipid-synthesizing and lipid-consuming tissues
and transport of fat-soluble vitamins, acyclic alcohols, β-carotene
4.
Phosphoproteins – contain a phosphate residue esterifying the –
OH of serine;
example: casein in milk
4.
Cofactor-proteins = protein + a nonprotein moiety.
The colored cofactor-proteins are chromoproteins:
–
Hemoproteins (contain heme)
–
Chorophyllo proteins (chlorophyll)
–
Cobamine proteins (vitamin B12)
–
Retinal proteins (vitamin A – aldehyde)
–
Flavoproteins (flavins)
Hemeproteins are classified in:
–
Nonenzymic: hemoglobin, myoglobin
–
Enzymic: cytochromes, catalases, peroxidases
The prosthetic group (non-protein component) = heme a metalloporphyrin
complex
Hemoglobin
Globular protein in the erythrocytes with molecular mass of
66,000-68,000
Structure: primary structure:
protein moiety (globin) + prosthetic group (heme)
1.The globin is an oligomer formed of 4 polypeptide chains in 2
subunits:
– 2 α chains containing
141 aminoacid radicals and
– 2 β chains containing
146 aminoacid radicals
– 2 α + 2 β → α2β2 → 2 αβ
The secondary structure – 8 α-helical segments (lettered A-H);
– the polar (hydrophylic) residues tend to be on the outside of the molecule,
– almost all the nonpolar (hydrophobic) residues are on the inside of the molecule
The tertiary structure: inside each subunit there is a hydrophobic pocket in which
one heme is held due to van der Waals bonds and ionic bonds
2. The heme is a heterocyclic molecule composed of
– protoporphyrin group = tetrapyrrole = four pyrrole groups linked by
methene bridges (–CH= );
protoporphyrin IX possesses substituents:
– methyl groups (-CH3)
at positions 1, 3, 5, 8
– vinyl (-CH=CH2)
at positions 2, 4
– propionyl groups (-CH2-CH2-COOH) at positions 6, 7
– Fe2+
His
HC CH 2
H3C
CH 3
2
3
4
1
N
N
N
2+
N
N
N
Fe2+
Fe2+
H3C
His
CH CH 2
N
N
5
8
6
7
H2C
H2C
H
H
O
H2C
H2C
COOH
COOH
His
CH 3
H
H
O
His
Fe2+ is bound in the center with 6 bonds:
– 4 bonds with the N of the tetrapyrole
ring,
– 1 bond with the proximal hystidine in
the F8 segment of the globin and
– 1 coordination bond free for binding
oxygen, on the other side of the
heme plane; close to bond 6 there is
a distal hystidine that influences the
interaction of heme with other ligands
Hemoglobin - Types
Normal:
Primitive (embryonal):
Hb P (Gower 1, Gower 2)
(disappears in 3 month)
Fetal:
Hb F α2γ2 = 70% of the Hb at birth moment
Adult:
Hb A α2β2
Hb A2 α2δ2
Hb A3 structurally changed β-chain, in old RBCs
In the adult blood: 95-96% HbA, 2-3% HbA2, 0.1-2% HbF
HbA2 and Hb F have a higher affinity for oxygen
Abnormal Hb:
Hb H
β4,
Bart’s Hb γ4,
Hb S (Sickle-cell Hb) glutamic acid in position 6 of β-chain is
changed with valine
His
Hemoglobin - Functions
N
N
Binds the oxygen and transfer it from the lungs to the tissues
Fe2+
Hb
+ 4 O2
 Hb(O2) 4 + 4 H2O
N
N
deoxyhemoglobin
oxyhemoglobin
O2
T-form (tense)
R-form (relaxed)
The process is dependent on pO2, pH, [CO2], [2,3-bisphosphoglycerate]
The first oxygen molecule becomes bound to the heme iron of α-chain which is
pulled into the porphyrin ring plane which results in the displacement of proximal
His. This detemines the rearrangement of the bonds with the other aminoacid
radical in the same subunit and a rupture of some of the ionic bonds between the
chains.
This facilitates the access of the second molecule of oxygen to the heme iron of
the α-chain. The addition of the second molecule of oxygen ruptures other ionic
bonds between the subunits.
The 3rd and 4th molecules of oxygen break the remaining ionic bonds. Thus the
quaternary structure is changed from T-form (tense) to R-form (relaxed). The Tform Hb affinity for the oxygen is 300 time lower than of the R-form.
The gradual increase of the Hb affinity for the oxygen has a sigmoid shape to the
oxygen binding curve and demonstrates the cooperative behavior of hemes
Hemoglobin - Derivatives
Carbhemoglobin: in interaction with CO2, this is added
to the globin -NH2
Hb-NH2 + CO2  Hb-NH-COO- + H+
carbhemoglobin
His
N
N
Fe2+
Carboxyhemoglobin (Hb-CO): Hb has an affinity 25,000
times greater for CO than for CO2; it cannot transfer O2
N
Methemoglobin: is formed by the action of oxidants
(nitrites, peroxides, ferricyanides, quinones); the Fe3+
can bind neither O2, nor CO2 inducing anoxia
N
N
CO
His
N
Fe3+
N
N
OH- (CN -)
Sulfhemoglobin: formed by irreversible reaction with
hydrogen sulphide, sulfonamides, aromatic amines
His
Chlorhemine: Teichman crystals with species specific
microscopic appearance (used in forensic laboratory)
N
N
Fe3+
N
N
Cl
Myoglobin
1 single-chain globin + 1 heme
The affinity for oxygen is 5 fold higher than the one of Hb
The curve of saturation with oxygen is a hyperbola
Heme-enzymes
Cytochromes a, b, c, d
– Cannot bind oxygen except Cyt a3
– Transfer of electrons as part in the respiratory chain of
mitochondria
-eCyt (Fe2+)
Cyt (Fe3+)
+eCatalases and peroxydases
– Take part in the decomposition of hydrogen peroxide
catalase
H2O2 + H2O2
O2 + 2 H2O
peroxydase
S-H2 + H2O2
S + 2 H2O