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GLOBULAR
HEMOPROTEINS
• Hemeproteins are a group of specialized proteins that
contain heme as a tightly bound prosthetic group.
• Heme is a complex of protoporphyrin IX and ferrous iron
(Fe2+) .
• The iron is held in the center of the heme molecule by
bonds to the four nitrogens of the porphyrin ring.
• The heme Fe2+ can form two additional bonds, one on
each side of the planar porphyrin ring.
• In myoglobin and hemoglobin, one of these positions is
coordinated to the side chain of a histidine residue of the
globin molecule, whereas the other position is available to
bind oxygen
A .Hemeprotein (cytochrome c(
B .Structure of heme
Myoglobin
Structure and function:
• Myoglobin, a hemeprotein present in heart and skeletal
muscle, functions both as a reservoir for oxygen, and as an
oxygen carrier that increases the rate of transport of oxygen
within the muscle cell.
• Myoglobin consists of a single polypeptide chain that is
structurally similar to the individual subunit polypeptide
chains of the hemoglobin molecule.
• Myoglobin is a compact molecule, with approximately 80 %
of its polypeptide chain folded into 8 stretches of α-helix.
These α-helical regions, labeled A to H
• The interior of the myoglobin molecule is composed almost
entirely of nonpolar amino acids.
• In contrast, polar amino acids are located almost
exclusively on the surface of the molecule.
• The heme group of myoglobin is located in a crevice in the
molecule between helix E and helix F, which is lined with
nonpolar amino acids. Notable exceptions are two histidine
residues.
A. Model of myoglobin showing helices A to H.
B. Schematic diagram of the oxygen-binding site of myoglobin
• One, the proximal histidine (F8), binds directly to the iron
of heme.
• The second, or distal histidine (E7), does not directly
interact with the heme group, but helps stabilize the
binding of oxygen to the ferrous iron.
• The protein, or globin, portion of myoglobin prevents the
oxidation of iron of heme.
Hemoglobin
Structure and function:
• Hemoglobin is found exclusively in red blood cells, where
its main function is to transport oxygen from the lungs to
the capillaries of the tissues.
• Hemoglobin A, the major hemoglobin in adults, is
composed of four polypeptide chains - two α chains and
two β chains - held together by noncovalent interactions.
• Each subunit has stretches of α-helical structure, and a
heme-binding pocket similar to that described for
myoglobin.
• However, the tetrameric hemoglobin molecule is
structurally and functionally more complex than
myoglobin.
• For example, hemoglobin can transport H+ and CO2 from
the tissues to the lungs, and can carry four molecules of
O2 from the lungs to the cells of the body.
• Furthermore, the oxygen-binding properties of
hemoglobin are regulated by interaction with allosteric
effectors .
3–6 %
Normal adult human hemoglobins.
[Note: The α-chains in these hemo-globins are identical]
HbA1c could be used as a monitor for the control of the blood
glucose level during the last 2 months for diabetic patients
Schematic diagram showing structural changes resulting
from oxygenation and deoxygenation of hemoglobin .
Quaternary structure of hemoglobin:
• The hemoglobin tetramer can be envisioned as being
composed of two identical dimers, (αβ(1 and (αβ(2, in
which the numbers refer to dimers one and two.
• The two polypeptide chains within each dimer are held
tightly together, primarily by hydrophobic interactions
• In contrast, the two dimers are able to move with
respect to each other, being held together primarily by
polar bonds.
• The weaker interactions between these mobile dimers
result in the two dimers occupying different relative
positions in deoxyhemoglobin as compared with
oxyhemoglobin
• T form: The deoxy form of hemoglobin is called the “T,” or
taut (tense) form.
•In the T form, the two αβ dimers interact through a network
of ionic bonds that constrain the movement of the
polypeptide chains. The T form is the low-oxygen-affinity
form of hemoglobin.
• R form: The binding of oxygen to hemoglobin causes the
rupture of some of the ionic bonds between the αβ dimers.
•This leads to a structure called the “R,” or relaxed form, in
which the polypeptide chains have more freedom of
movement . The R form is the high- oxygen-affinity form of
hemoglobin.
Binding of oxygen to myoglobin and hemoglobin
• Myoglobin can bind only one molecule of oxygen (O2),
because it contains only one heme group.
• In contrast, hemoglobin can bind four oxygen
molecules—one at each of its four heme groups.
• The degree of saturation (Y) of these oxygen-binding
sites on all myoglobin or hemoglobin molecules can vary
between zero (all sites are empty) and 100% (all sites
are full
Oxygen dissociation curves for myoglobin and hemoglobin
Oxygen dissociation curve:
• A plot of Y measured at different partial pressures of
oxygen (pO2) is called the oxygen dissociation curve.
• The curves for myoglobin and hemoglobin show
important differences.
• This graph illustrates that myoglobin has a higher
oxygen affinity at all pO2 values than does hemoglobin
Allosteric effects:
• The ability of hemoglobin to reversibly bind oxygen is
affected by the pO2 (through heme-heme interactions ,
the pH of the environment, the pCO2, the availability of
2,3-bisphosphoglycerate and CO.
• These are collectively called allosteric (“other site”)
effectors, because their interaction at one site on the
hemoglobin molecule affects the binding of oxygen to
heme groups at other locations on the molecule.
• [Note: The binding of oxygen to myoglobin is not
influenced by allosteric effectors.]
 Effect of O2 binding (Heme-heme interactions):
 The sigmoidal oxygen-binding curve reflects specific
structural changes that are initiated at one heme group and
transmitted to other heme groups in the hemoglobin
tetramer.
 The net effect is that the affinity of hemoglobin for the last
oxygen bound is approximately 300 times greater than its
affinity for the first oxygen bound.
 O2 favors the R- form of Hb.
 Deoxygenation favors the T- form of Hb.
Effect of pH and CO2 (Bohr effect):
• The release of oxygen from hemoglobin is enhanced when the pH is
lowered ( H+) or when the hemoglobin is in the presence of an
increased partial pressure of CO2.
• Both result in a decreased oxygen affinity of hemoglobin and both,
stabilize the T state.
• In the tissues, CO2 is converted by carbonic anhydrase (CA) to
carbonic acid:
CO2 + H2O
H2CO3
which spontaneously loses a proton, becoming bicarbonate
(the major blood buffer):
H2CO3
HCO3¯ + H+
• The H+ produced by this pair of reactions   pH
• H+ increases the ionic bonds on Hb T-form  O2 release
to the tissues.
• Lactic acid produced during muscular exercise   pH
Effect of 2,3-bisphosphoglycerate on oxygen affinity:
• 2,3- Bisphosphoglycerate (2,3-BPG) is an important regulator of the
binding of oxygen to hemoglobin.
• It is the most abundant organic phosphate in the red blood cell, where
its concentration is approximately that of hemoglobin.
• 2,3-BPG is synthesized from an intermediate of the glycolysis.
Binding of 2,3-BPG to deoxyhemoglobin:
• 2,3-BPG decreases the oxygen affinity of hemoglobin by binding to
deoxyhemoglobin but not to oxyhemoglobin.
• This preferential binding stabilizes the T conformation of
deoxyhemoglobin.
Response of 2,3-BPG levels to chronic hypoxia or anemia:

The concentration of 2,3-BPG in the red blood cell increases in response
to chronic hypoxia or anemia.
• Elevated 2,3-BPG levels lower the oxygen affinity of hemoglobin,
permitting greater unloading of oxygen in the capillaries of the tissues.
Binding of CO:
• Carbon monoxide (CO) binds tightly (but reversibly) to
the hemoglobin iron, forming carboxyhemoglobin.
• When carbon monoxide binds to one or more of the four
heme sites, hemoglobin shifts to the relaxed
conformation (R-form), causing the remaining heme sites
to bind oxygen with high affinity.
• As a result, the affected hemoglobin is unable to release
oxygen to the tissues.
• [Note: The affinity of hemoglobin for CO is 220 times
greater than for oxygen.
Factors favoring the T-form of Hb. are:
• Deoxygenation
• Low pH (  H+)
• CO2
• Lactic acid
• 2,3 bisphosphoglycerate
Factors favoring the R-form of Hb. are:
• O2
• CO
Hemoglobinopathies:
• Hemoglobinopathies have traditionally been defined as a
family of genetic disorders caused by production of a
structurally abnormal hemoglobin molecule, synthesis of
insufficient quantities of normal hemoglobin, or, rarely,
both.
• Sickle cell anemia (Hb S), hemoglobin C disease (Hb C),
and the thalassemia syndromes are representative
hemoglobinopathies that can have severe clinical
consequences.
• The first two conditions result from production of
hemoglobin with an altered amino acid sequence
(qualitative hemoglobinopathy), whereas the
thalassemias are caused by decreased production of
normal hemoglobin (quantitative hemoglobinopathy).
Sickle cell disease (hemoglobin S disease)
(SCD)
• Sickle cell disease (also called sickle cell anemia) is a
genetic disorder of the blood caused by a single
nucleotide alteration (a point mutation( in the β-globin
gene.
• Sickle cell disease is an autosomal recessive disorder.
• It occurs in individuals who have inherited two mutant
genes (one from each parent) that code for synthesis of
the β chains of the globin molecules.
 A molecule of Hb S contains two normal α-globin chains
and two mutant β-globin chains (βS(, in which glutamate
at position six has been replaced with valine .
 Therefore, during electrophoresis at alkaline pH, Hb S
migrates more slowly toward the anode (positive electrode)
than does Hb A .
 Electrophoresis of hemoglobin obtained from lysed red
blood cells is routinely used in the diagnosis of sickle cell
trait and sickle cell disease.
 HbS is slower in electrophoretic motility than HbA at pH 8.6
 The lifetime of an erythrocyte in SCD is less than 20 days
compared to 120 days in normal RBC, hence causing
anemia.
 HbS is less soluble than HbA and precipitates specially
when in T-form giving the RBCs the sickle shape.