Contribution of the gamma-carboxyl group of Glu-43(beta) to the alkaline Bohr effect of hemoglobin A.
Glu-43(beta) of hemoglobin A exhibits a high degree of chemical reactivity around neutral pH for amidation with nucleophiles in the presence of carbodiimide. Such a reactivity is unusual for the side-chain carboxyl groups of proteins. In addition, the reactivity of Glu-43(beta) is also sensitive to the ligation state of the protein [Rao, M. J., & Acharya, A. S. (1991) J. Protein Chem. 10, 129-138]. The influence of deoxygenation of hemoglobin A on the chemical reactivity of the gamma-carboxyl group of Glu-43(beta) has now been investigated as a function of pH (from 5.5 to 7.5). The chemical reactivity of Glu-43(beta) for amidation increases upon deoxygenation only when the modification reaction is carried out above pH 6.0. The pH-chemical reactivity profile of the amidation of hemoglobin A in the deoxy conformation reflects an apparent pKa of 7.0 for the gamma-carboxyl group of Glu-43(beta). This pKa is considerably higher than the pKa of 6.35 for the oxy conformation. The deoxy conformational transition mediated increase in the pKa of the gamma-carboxyl group of Glu-43(beta) implicates this carboxyl group as an alkaline Bohr group. The amidated derivative of hemoglobin A with 2 mol of glycine ethyl ester covalently bound to the protein was isolated by CM-cellulose chromatography.(ABSTRACT TRUNCATED AT 250 WORDS)
Rao
M J
MJ
Department of Medicine, Albert Einstein College of Medicine, Bronx, New York 10461.
Acharya
A S
AS
D000595:Amino Acid Sequence; D002848:Chromatography, DEAE-Cellulose; D005971:Glutamates; D018698:Glutamic Acid; D006441:Hemoglobin A / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006451:Hemoglobin, Sickle / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D046911:Macromolecular Substances; D010108:Oxyhemoglobins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*
D005971:Glutamates; D006451:Hemoglobin, Sickle; D046911:Macromolecular Substances; D010108:Oxyhemoglobins; D018698:Glutamic Acid; D006441:Hemoglobin A; C029774:oxyhemoglobin A