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Myoglobin
Hemoglobin
Role of 2,3-BPG

Dr. Paul Reisberg's excellent pages on
Hemoglobin
Myoglobin

These pages have wonderful Protein Database images that one can manipulate to view various substituents such as the hemes and 2,3-BPG



Myoglobin

Myoglobin and hemoglobin are hemeproteins whose physiological importance is principally related to their ability to bind molecular oxygen. Myoglobin is a monomeric heme protein found mainly in muscle tissue where it serves as an intracellular storage site for oxygen. During periods of oxygen deprivation oxymyoglobin releases its bound oxygen which is then used for metabolic purposes.
The tertiary structure of myoglobin is that of a typical water soluble globular protein. Its secondary structure is unusual in that it contains a very high proportion (75%) of a-helical secondary structure. A myoglobin polypeptide is comprised of 8 separate right handed a-helices, designated A through H, that are connected by short non helical regions. Amino acid R-groups packed into the interior of the molecule are predominantly hydrophobic in character while those exposed on the surface of the molecule are generally hydrophilic, thus making the molecule relatively water soluble.
Each myoglobin molecule contains one heme prosthetic group inserted into a hydrophobic cleft in the protein. Each heme residue contains one central coordinately bound iron atom that is normally in the Fe2+, or ferrous, oxidation state. The oxygen carried by hemeproteins is bound directly to the ferrous iron atom of the heme prosthetic group. Oxidation of the iron to the Fe3+, ferric, oxidation state renders the molecule incapable of normal oxygen binding. Hydrophobic interactions between the tetrapyrrole ring and hydrophobic amino acid R groups on the interior of the cleft in the protein strongly stabilize the heme protein conjugate. In addition a nitrogen atom from a histidine R group located above the plane of the heme ring is coordinated with the iron atom further stabilizing the interaction between the heme and the protein. In oxymyoglobin the remaining bonding site on the iron atom (the 6th coordinate position) is occupied by the oxygen, whose binding is stabilized by a second histidine residue.
Carbon monoxide also binds coordinately to heme iron atoms in a manner similar to that of oxygen, but the binding of carbon monoxide to heme is much stronger than that of oxygen. The preferential binding of carbon monoxide to heme iron is largely responsible for the asphyxiation that results from carbon monoxide poisoning.
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Hemoglobin

Hemoglobin is an [a(2):b(2)] tetrameric hemeprotein found in erythrocytes where it is responsible for binding oxygen in the lung and transporting the bound oxygen throughout the body where it is used in aerobic metabolic pathways. Each subunit of a hemoglobin tetramer has a heme prosthetic group identical to that described for myoglobin. The common peptide subunits are designated a, b, g and d which are arranged into the most commonly occurring functional hemoglobins.
Although the secondary and tertiary structure of various hemoglobin subunits are similar, reflecting extensive homology in amino acid composition, the variations in amino acid composition that do exist impart marked differences in hemoglobin's oxygen carrying properties. In addition, the quaternary structure of hemoglobin leads to physiologically important allosteric interactions between the subunits, a property lacking in monomeric myoglobin which is otherwise very similar to the a-subunit of hemoglobin.
Comparison of the oxygen binding properties of myoglobin and hemoglobin (see this link from Thomas J. Herbert) illustrate the allosteric properties of hemoglobin that results from its quaternary structure and differentiate hemoglobin's oxygen binding properties from that of myoglobin. The curve of oxygen binding to hemoglobin is sigmoidal typical of allosteric proteins in which the substrate, in this case oxygen, is a positive homotropic effector. When oxygen binds to the first subunit of deoxyhemoglobin it increases the affinity of the remaining subunits for oxygen. As additional oxygen is bound to the second and third subunits oxygen binding is further, incrementally, strengthened, so that at the oxygen tension in lung alveoli, hemoglobin is fully saturated with oxygen. As oxyhemoglobin circulates to deoxygenated tissue, oxygen is incrementally unloaded and the affinity of hemoglobin for oxygen is reduced. Thus at the lowest oxygen tensions found in very active tissues the binding affinity of hemoglobin for oxygen is very low allowing maximal delivery of oxygen to the tissue. In contrast the oxygen binding curve for myoglobin is hyperbolic in character indicating the absence of allosteric interactions in this process.
The allosteric oxygen binding properties of hemoglobin arise directly from the interaction of oxygen with the iron atom of the heme prosthetic groups and the resultant effects of these interactions on the quaternary structure of the protein. When oxygen binds to an iron atom of deoxyhemoglobin it pulls the iron atom into the plane of the heme. Since the iron is also bound to histidine F8, this residue is also pulled toward the plane of the heme ring. The conformational change at histidine F8 is transmitted throughout the peptide backbone resulting in a significant change in tertiary structure of the entire subunit. Conformational changes at the subunit surface lead to a new set of binding interactions between adjacent subunits. The latter changes include disruption of salt bridges and formation of new hydrogen bonds and new hydrophobic interactions, all of which contribute to the new quaternary structure.
The latter changes in subunit interaction are transmitted, from the surface, to the heme binding pocket of a second deoxy subunit and result in easier access of oxygen to the iron atom of the second heme and thus a greater affinity of the hemoglobin molecule for a second oxygen molecule. The tertiary configuration of low affinity, deoxygenated hemoglobin (Hb) is known as the taut (T) state. Conversely, the quaternary structure of the fully oxygenated high affinity form of hemoglobin (HbO2) is known as the relaxed (R) state.

If using a Javascript supporting browser one can see the T to R transition at this site.

In addition to transporting oxygen from lungs to peripheral tissues, hemoglobin molecules play an important role in transporting CO2 in the opposite direction. N-terminal amino groups of the T form of hemoglobin are available for reaction with CO2 and about 15% of the CO2 formed in tissues is carried to the lung covalently bound to the N-terminal nitrogens as carbamate:

CO2 + Hb-NH2 <-----> H+ + Hb-NH-COO-

In the lung the high oxygen partial pressure favors formation of the R form resulting in reversal of the carbamate with release and exhalation of the CO2.
During the conversion from T form to R form several R groups on the surface of hemoglobin subunits also become available to dissociate protons:

4O2 + Hb <--------> nH+ + Hb(O2)4

The above two equations illustrate that the conformation of hemoglobin and its oxygen binding are sensitive to hydrogen ion concentration. These effects of hydrogen ion concentration are responsible for the well known Bohr effect in which increases in hydrogen ion concentration decrease the amount of oxygen bound by hemoglobin at any oxygen concentration (partial pressure).

Representation of the transport of CO2 from the tissues to the blood with delivery of O2 to the tissues. The opposite process occurs when O2 is taken up from the alveoli of the lungs and the CO2 is expelled. All of the processes of the transport of CO2 and O2 are not shown such as the formation and ionization of carbonic acid in the plasma. The latter is a major mechanism for the transport of CO2 to the lungs, i.e. in the plasma as HCO3-. The H+ produced in the plasma by the ionization of carbonic acid is buffered by phosphate (HPO42-) and by proteins. Additionally, some 15% of the CO2 is transported from the tissues to the lungs as hemoglobin carbamate.

Tissue CO2 is also carried to the lungs as the dissolved gas and as bicarbonate formed spontaneously and by the enzyme carbonic anhydrase which converts CO2 and H2O to carbonic acid. The carbonic acid thus formed spontaneously ionizes to proton and bicarbonate ion:

CO2 + H2O --------> H2CO3 ------> H+ + HCO3-

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Role of 2,3-bisphosphoglycerate (2,3-BPG)

The compound 2,3-bisphosphoglycerate (2,3-BPG), derived from the glycolytic intermediate 1,3-bisphosphoglycerate, is a potent allosteric effector on the oxygen binding properties of hemoglobin.
The formation of 2,3-BPG is diagrammed. In the deoxygenated T conformer, a cavity capable of binding 2,3-BPG forms in the center of the molecule. 2,3-BPG can occupy this cavity stabilizing the T state. Conversely, when 2,3-BPG is not available, or not bound in the central cavity, Hb can be converted to HbO2 more readily. Thus, like increased hydrogen ion concentration, increased 2,3-BPG concentration favors conversion of R form Hb to T form Hb and decreases the amount of oxygen bound by Hb at any oxygen concentration. Hemoglobin molecules differing in subunit composition are known to have different 2,3-BPG binding properties with correspondingly different allosteric responses to 2,3-BPG. For example, HbF (the fetal form of hemoglobin) binds 2,3-BPG much less avidly than HbA (the adult form of hemoglobin) with the result that HbF in fetuses of pregnant women binds oxygen with greater affinity than the mothers HbA, thus giving the fetus preferential access to oxygen carried by the mothers circulatory system.
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This article has been modified by Dr. M. Javed Abbas.
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20:37 21/12/2002