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BIOCHEMISTRY | PROTEIN STRUCTURE AND FUNCTION | GLOBULAR PROTEIN | NOTED BY FAKHRY (IG @SFAKHRYM)

v Hemoglobin VS Myoglobin

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BIOCHEMISTRY | PROTEIN STRUCTURE AND FUNCTION | GLOBULAR PROTEIN | NOTED BY FAKHRY (IG @SFAKHRYM)

v GLOBULAR HEMEPROTEINS
o They are a group of specialized proteins that contain heme as a tightly bound prosthetic group
...

•
• The iron is held in the center of the heme molecule by bonds to the four nitrogens of the porphyrin
• The heme Fe2+ can form two additional bonds, one on each side of the planar porphyrin ring
o

Myoglobin structure and function
• It is a hemeprotein present in heart and skeletal muscle, function as an oxygen reservoir and as an oxygen carrier
• It consists of a single polypeptide chain that is structurally similar to the individual polypeptide chains of the tetrameric hemoglobin
molecule
α-helical content
•
§ Myoglobin is a compact molecule, with ~80% of its polypeptide chain folded into eight stretches of α-helix
§ These α-helical regions are terminated either by the presence of proline, whose five-membered ring can not be accommodated in
an α-helix, or by β-bends and loops stabilized by hydrogen bonds and ionic bonds
• Location of polar and nonpolar amino acid residues
§ The interior of the globular myoglobin molecule is composed almost entirely of nonpolar aas
• They are packed closely together forming a structure stabilized by hydrophobic interactions between these clustered
residues
§ In contrast, polar aas are located almost exclusively on the surface, where they can not form hydrogen bond, both with each other
and with water
• Binding of the heme group
§ The heme group of the myoglobin molecule sits in a crevice, which is lined with nonpolar amino acids
Exceptions of two histidine residues
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BIOCHEMISTRY | PROTEIN STRUCTURE AND FUNCTION | GLOBULAR PROTEIN | NOTED BY FAKHRY (IG @SFAKHRYM)
o

Hemoglobin structure and function
• Found exclusively in RBC
• Hemoglobin A, the major hemoglobin in adults, composed of four polypeptide chains (two α-chains and two β-chains) held together by
noncovalent interactions
• Each chain (subunit) has stretches of α-helical structure and a hydrophobic heme-binding pocket similar to that described for myoglobin
...
The two polypeptide chains
within each dimer are held tightly together primarily by hydrophobic interactions (in this instance, hydrophobic amino acid
residues are localized not only in the interior of the molecule but also in a region on the surface of each subunit)
§ In contrast, the two dimers are held together primarily by polar bonds
§ T form (taut (tense) form)
• It is the deoxy form of hemoglobin; it is the low-oxygen-affinity form of hemoglobin
• In T form, the two αβ dimers interact through a network of ionic bonds and hydrogen bond that constrain the movement
of the polypeptide chains
§ R form (relaxed form)
• The binding of oxygen to hemoglobin; It is the high-oxygen-affinity for of hemoglobin

o

Oxygen binding to myoglobin and hemoglobin
• Myoglobin can bind only one molecule of O2, because it contains only one heme group
• Hemoglobin can bind four molecules of O2, 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 curve: it is a plot of %saturation of with oxygen measured at different partial pressures of oxygen (pO2)
§ In the graph, it illustrates that Hb has a higher oxygen at all pO2 values than does myoglobin
§ The partial pressure of oxygen needed to achieve half-saturation of the binding sites (P50) is approximately 1 mm Hg for myoglobin
and 26 mm Hg for Hemoglobin
• The higher the oxygen affinity, the lower the P50

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BIOCHEMISTRY | PROTEIN STRUCTURE AND FUNCTION | GLOBULAR PROTEIN | NOTED BY FAKHRY (IG @SFAKHRYM)
Myoglobin
• It is a hyperbolic shape
• It reflects the fact that myoglobin reversibly binds a single molecule of oxygen
• Equation: Mb + O2 <--> MbO2
• Myoglobin is designed to bind oxygen released by hemoglobin at the low pO2 found in muscle, Myoglobin, in turn,
releases oxygen within the muscle cell in response to oxygen demand
• Hemoglobin
• It is a sigmoidal shape
• Indicating that the subunits cooperate in binding oxygen
• Cooperative binding of Oxygen by the four subunits of hemoglobin means that the binding of an oxygen molecule
at one heme group (initiated at one heme group) increases the oxygen affinity of the remaining heme groups in
the same hemoglobin tetramer (referred to heme-heme interaction)
• Although it is more difficult for the first oxygen molecule to bind to hemoglobin, the subsequent binding
of oxygen occurs with high affinity
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• Example: In the lung, the concentration of O2 is high, and hemoglobin becomes virtually saturated ("loaded") with O2
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§ Mechanism of the Bohr effect
• This effect reflects the fact that the deoxy form of Hb has greater affinity for protons that does oxyhemoglobin
This effect is caused by ionizable groups such as specific histidine side chains that have a higher pKa in
•
deoxyhemoglobin
• Therefore, an increase in the concentration of protons (resulting in a decreases in pH) causes these groups
to become protonated (charged) and able to form ionic bonds (salt bridges)
• The bonds preferentially stabilize the deoxy form of Hb, producing a decrease in oxygen affinity
[Oxyhemoglobin]
HbO2 + H+ <--> HbH + O2 [deoxy]
•
• Where an increase in protons (or lower pO2) shifts the equilibrium to the right (favoring deoxyhemoglobin),
whereas an increase in pO2 (or a decrease in protons) sifts the equilibrium to the left

Effect of 2,3-bisphosphoglycerate (2,3-BPG) on oxygen affinity
§ It is an important regulator of the binding of oxygen to Hb
§ It is the most abundant organic phosphate in the RBC; it is synthesized from an intermediate of glycolytic pathway

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BIOCHEMISTRY | PROTEIN STRUCTURE AND FUNCTION | GLOBULAR PROTEIN | NOTED BY FAKHRY (IG @SFAKHRYM)
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Binding of 2,3-BPG to deoxyhemoglobin
• The binding decreases the O2 affinity of Hb by binding to deoxyhemoglobin
• HbO2 + 2,3-BPG <--> Hb-2,3-BPG + O2
Binding site of 2,3-BPG
• One molecule of 2,3-BPG binds to a pocket, formed by the two β-globin chains, in the center of the deoxyhemoglobin tetramer
• This pocket contains several positively charged amino acids that form ionic bonds with the negatively charged phosphate
groups of 2,3-BPG
Shift of the oxygen dissociation curve
• In RBC, the presence of 2,3-BPG significantly reduces the affinity of hemoglobin for oxygen, shifting the oxygen-dissociation curve
to the right
• This reduced affinity enables hemoglobin to release oxygen efficiently at the partial pressures found in the tissues
Response of 2,3-BPG levels to chronic hypoxia or anemia
• In chronic hypoxia, such as COPD like emphysema, or at high altitudes; where circulating hemoglobin may have difficulty receiving
receiving sufficient oxygen
...
However, severely ill patients may be
compromised if transfused with large quantities of such 2,3-BPG-"stripped" blood

Binding of CO2
§ Most of the CO2 produced in metabolism is hydrated and transported as bicarbonate ion
• However, some CO2 is carried as carbamate bound to the N-terminal amino groups of hemoglobin (forming carbaminohemoglobin)
Hb - NH2 + CO2 <--> Hb - NH - COO- + H+

§
§

The binding of CO2 stabilizes the T or deoxy form of hemoglobin, resulting in a decrease in its affinity for oxygen and a right shift in the oxygendissociation curve
In the lungs, CO2 dissociates from the Hb and is released in the breath

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BIOCHEMISTRY | PROTEIN STRUCTURE AND FUNCTION | GLOBULAR PROTEIN | NOTED BY FAKHRY (IG @SFAKHRYM)
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Binding of CO
§ CO binds tightly (but reversibly) to the hemoglobin iron, forming carboxyhemoglobin
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Result, the
affected hemoglobin is unable to release oxygen to the tissues
§ The affinity of Hb for CO is 220 times greater than oxygen
§ CO poisoning is treated with 100% oxygen at high pressure (hyperbaric oxygen therapy), which facilitates the dissociation of CO from Hb
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• The γ chains are members of the β-globin gene family
§ HbF synthesis during development
• First month after conception, embryonic hemoglobins such as Hb Gower 1, composed of two α-like zeta (ζ) chains and two
β-like epsilon (ε) chains (ζ2ε2) --> are synthesized by embryonic yolk sac
• Fifth week of gestation, the site of globin synthesis shifts, first to the liver and then to the marrow, and the primary
product is HbF
• HbF is the major hemoglobin found in the fetus and newborn (about 60% of the total hemoglobin in the RBC
during the last months of fetal life)
• HbA synthesis starts in the bone marrow at about the eighth month of pregnancy and gradually replaces HbF
• Note: HbF represents less than 1% of the hemoglobin in most adults and is concentrated in RBC known as F cells
§ Binding of 2,3-BPG to HbF
• Under physiologic conditions, HbF has a higher affinity for oxygen than does HbA as a result of HbF only weakly binding
2,3-BPG
• Note: the γ-globin chains of HbF lack some of the positively charged amino acids that are responsible for binding 2,3-BPG
in the β-globin chains

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BIOCHEMISTRY | PROTEIN STRUCTURE AND FUNCTION | GLOBULAR PROTEIN | NOTED BY FAKHRY (IG @SFAKHRYM)
Because 2,3-BPG serves to reduce the affinity of hemoglobin for oxygen, the weaker interaction between 2,3-BPG
and HbF results in a higher oxygen affinity for HbF relative to HbA; in contrast, if both HbA and HbF are stripped of
their 2,3-BPG, they then have a similar affinity for oxygen
The higher oxygen affinity of HbF facilitates the transfer of oxygen from the maternal circulation across the placenta to
the RBC of the fetus
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Hemoglobin A2
§ It is a minor component of normal adult Hb
§ First appearing shortly before birth and, ultimately, consisting about 2% of the total hemoglobin
§ It is composed of two α-globin chains and two δ-globin chains (α2δ2)
Hemoglobin A1c
§ HbA1c is the most abundant form of glycosylated Hb
§ Under physiologic conditions, HbA is slowly and nonenzymatically glycosylated (glycated);
• The extent of glycosylation being dependent on the plasma concentration of a particular hexose
§ HbA1c has glucose residues attached predominantly to the NH2 groups of the N-terminal valines of the β-globin chains
§ Increased amount of HbA1c is found in RBC of patients with Diabetes Mellitus, because their HbA has contact with higher glucose
concentrations during the 120-day lifetime of these cells

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BIOCHEMISTRY | PROTEIN STRUCTURE AND FUNCTION | GLOBULAR PROTEIN | NOTED BY FAKHRY (IG @SFAKHRYM)
v ORGANIZATION OF THE GLOBIN GENES
o α-Gene family
• The genes coding for the α-globin and β-globin subunits occur in two separates gene clusters (or families) located on two different

o

o

chromosomes
§ The α-gene cluster on chromosome 16 contains two genes for the α-globin chains
• It also contains the ζ gene that is expressed early in development as an α-globin-like component of embryonic
hemoglobin
• Note: globin gene families also contain globin-like genes that are not expressed, that is, their genetic information is not used to
produce globin chains - pseudogens
β-Gene family
• A single gene for the β-globin chain is located on chromosome 11
• Additional four β-globin like genes: the ε gene (which, like the ζ gene, is expressed early in embryonic development) , two γ genes
(Gγ and Aγ that are expressed in HbF), and the δ gene that code for the globin chain found in the minor adult hemoglobin HbA2
Steps in globin chain synthesis
• Expression of a globin gene begins in the nucleus of RBC precursors, where the DNA sequence encoding the gene is transcribed

v HEMOGLOBINOPATHY
o It is a group of genetic disorders caused by production of a structurally abnormal hemoglobin molecule
o Representative hemoglobinopathies are Sickle cell anemia (HbS), hemoglobin C disease (HbC), hemoglobin SC disease (HbS + HbC = HbSC),

and the Thalassemias
• The first three results from production of hemoglobin with an altered amino acid sequence (qualitative hemoglobinopathy);
thalassemias are cause by decreased production of normal hemoglobin (quantitative hemoglobinopathy)

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