Hemoglobin, a chromo protein, found exclusively in red blood cells is actually a conjugated protein containing heme as prosthetic group and globin as the protein part apoprotein.
The normal concentration of Hb in an adult varies from 14.0 to 16.0 gm%. Approximately 90 mg/kg of Hb is produced and destroyed in the body every day. Hb has a molecular weight of about 67,000. Each gram of Hb contains 3.4 mg of iron.
Heme is present as a prosthetic group in hemoglobin as well as in myoglobin, cytochromes, peroxidases, catalases and tryptophan pyrrolases etc.
Heme is produced by the combination of iron with a porphyrin ring. The heme portion is alike in all forms of hemoglobin.
Structure of Heme
Heme is a derivative of porphyrin. Porphyrins are cyclic compounds formed by the fusion of 4 pyrrole rings linked by methenyl bridges (=CH-). Since an atom of iron is present heme is a ferroprotoporphyrin. These rings are names as I,II,III, IV and the bridges are names as Alpha, beta, gamma and delta (Figure-1). Porphyrins contain side chains attached to each of the other four pyrrole rings. Different Porphyrins vary in the nature of the side chains that are attached to each of the pyrrole rings.
Figure-1-Showing the structure of heme molecule
Heme consists of one ferrous atom (Fe++) that is co-ordinated in the centre of the tetra pyrrole ring of protoporphyrin IX (Figure-1). The double bonds are resonating and therefore keep shifting in their position. When the ferrous atom in heme gets oxidized to ferric form, Hematin is formed, which loses the property of carrying oxygen and is brown in color, as compared to that of heme which is red in color.
Structure of Globin
Different hemoglobins are produced during embryonic, fetal, and adult life . Each consists of a tetramer of globin polypeptide chains. The major adult hemoglobin, HbA, has the structure α2β2. HbF (α2Υ2) predominates during most of gestation, and HbA2 (α2δ2) is minor adult hemoglobin.
Each polypeptide chain contains heme in the heme pocket. Thus one Hb molecule contains 4 Heme units.
The subunits of hemoglobin are arranged in a tetrahedral array with a tight spherical overall appearance and each individual polypeptide is folded in such a manner to maximize polar residues being on the exposed surface and non-polar interactions being internal, making this large protein water-soluble. The interior surface of the molecule lined with nonpolar groups, forms a hydrophobic pocket into which heme is inserted.
Figure-2- showing heme pocket in each of the polypeptide chain of hemoglobin.
The arrangement of polypeptides is held together by hydrogen bonding, hydrophobic interactions and multiple ionic interactions, that take place at the contact points between subunits. These subunit interactions play a critical role in the binding of oxygen to hemoglobin.
In the amino acid sequence of each polypeptide chain, certain residues appear to be critical to stability and function. Such residues are usually the same (invariant) in α or β chains. The NH2-terminal valines of the beta chains are important in 2,3-BPG interactions (bisphosphoglycerate has replaced the older term diphosphoglycerate). The C-terminal residues are important in the salt bridges (See details below)
Each heme moiety can bind a single oxygen molecule; a molecule of hemoglobin can transport up to four oxygen molecules. Each heme unit holds an iron ion in such a way that the iron can interact with an oxygen molecule, forming oxyhemoglobin , Blood containing RBCs filled with oxyhemoglobin is bright red. The iron–oxygen interaction is very weak; the two can easily be separated without damaging the heme unit or the oxygen molecule. The binding of an oxygen molecule to the iron in a heme unit is therefore completely reversible. A hemoglobin molecule in which the iron has separated from the oxygen molecule is called deoxyhemoglobin . Blood containing RBCs filled with deoxyhemoglobin is dark red–almost burgundy.
Primary structure of hemoglobin
Normal alpha chain contains 141 AA residues in linear sequence. The non-α (β,Υ and δ) chains are all 146 amino acids in length; the beta chain begins with valine and histidine. The C-terminal residues are Tyr b145 and His b146. The delta chain (of hemoglobin A2) differs from the beta chain (of hemoglobin A) in only 10 residues. The first eight residues and the C-terminal residues (127 to 146) are the same in delta and beta chains. Tetramers of beta chains (hemoglobin H) may be found in a thalassemia.
The gamma chain of fetal hemoglobin (hemoglobin F) differs from the beta chain by 39 residues. The N-terminal residues of the gamma chain and beta chain are glycine and valine respectively, while the C-terminal residues, Tyr145 and His146, are the same as in gamma and beta chains. Appreciable quantities of free gamma chains are found in the red cells of some infants with a thalassemia; free gamma chains, like beta chains, can form homotetramers known as hemoglobin Bart’s.
The gamma genes are duplicated: one codes for glycine (Gg) and the other for alanine (Ag)7 at residue 176, giving rise to two kinds of gamma chains.
Secondary structure of hemoglobin
About 75 percent of the amino acids in α or β chains are in a helical arrangement. All studied hemoglobins have a similar helical content. Eight helical areas, lettered A to H, occur in the β chains. Hemoglobin nomenclature specifies that amino acids within helices are designated by the amino acid number and the helix letter, while amino acids between helices bear the number of the amino acid and the letters of the two helices. Thus, residue EF3 is the third residue of the segment connecting the E and F helices, while residue F8 is the eighth residue of the F helix. Alignment according to helical designation makes homology evident: residue F8 is the proximal heme-linked histidine, and the histidine on the distal side of the heme is E7.
Tertiary structure of α or β chains
The tertiary folding of each globin chain forms an approximate sphere. Tertiary folding gives rise to at least 3 functionally important characteristics of the hemoglobin molecule :
1- Polar or charged side chains tend to be directed to the outside surface of the subunit and, conversely, non-polar structures tend to be directed inwards. The effect of this is to make the surface of the molecule hydrophilic and the interior hydrophobic.
2- An open-toped cleft in the surface of the subunit known as haem pocket is created. This hydrophobic cleft protects the ferrous ion from oxidation.
3- The amino acids, which form the inter-subunit bonds responsible for maintaining the quaternary structure, and thus the function, of the haemoglobin molecule are brought into the correct orientation to permit these bonds to form.
The heme group is located in a crevice between the E and F helices in each chain). The highly polar propionate side chains of the heme are on the surface of the molecule and are ionized at physiologic pH. The rest of the heme is inside the molecule, surrounded by nonpolar residues except for two histidines. The iron atom is linked by a coordinate bond to the imidazole nitrogen (N) of histidine F8; the E7 distal histidine, on the other side of the heme plane, is not bonded to the iron atom but is very close to the ligand-binding site.
Quaternary structure of hemoglobin
The Hb tetramer can be envisioned as being composed of two identical dimers (αβ)1and (αβ)2, where the number refers to dimer 1 and 2.
Figure-3 showing the T and R forms of hemoglobin. T form is the taut structure while R form is the relaxed form
The two polypeptide chains within each dimer are held together tightly, primarily by hydrophobic and ionic interactions, also by hydrogen bonding. The two dimers on the other hand can move with respect to each other, being held together primarily by polar bonds.
Figure-4- showing the helical structure of globin chains forming heme pockets upon folding. The adult Hb has a compact globular, tetrameric structure.
The weaker interactions between these mobile dimers result in the two dimers occupying different relative positions in deoxyhemoglobin compared to oxyhemoglobin.
a) T form
The deoxy form of hemoglobin is called the “T” form or taut or tense form. In this form, the two αβ dimers interact through a network of ionic bonds and hydrogen bonds that constrain the movement of the polypeptide chains. The T form is the low oxygen affinity form of Hemoglobin (Figure-3)
b) R form
The binding of hemoglobin causes rupture of some of the ionic bonds and hydrogen bonds between the αβ dimers. This leads to a structure called “R” or relaxed form, in which the polypeptide chains have more freedom of movement. The R form is the high affinity form of hemoglobin (Figure-3)
Please help "Biochemistry for Medics" by CLICKING ON THE ADVERTISEMENTS above!