Trypsin-catalyzed synthesis of peptide bond in human hemoglobin. Oxygen binding characteristics of Gly-NH2(142 alpha) Hb.

Incubation of human hemoglobin (Hb) A with glycineamide (Gly-NH2) in the presence of trypsin gives rise to two bands of approximately equal amount on the isoelectric focusing gel. Structural analyses, including digestion with trypsin and carboxypeptidases A and B, have shown that one is unmodified Hb A and the other is Gly-NH2(142 alpha) Hb in which Gly-NH2 is bound to the COOH-terminal residue of the alpha subunit (Arg 141 alpha) via peptide bond. Gly-NH2(142 alpha) Hb exhibits slightly increased affinity for oxygen and slightly reduced heme-heme interaction as compared with normal Hb A. Although Gly-NH2(142 alpha) Hb exhibits normal 2,3-diphosphoglycerate effect, the alkaline Bohr effect and chloride effect are diminished. These oxygen binding properties are attributable to abolished binding of chloride to the anion binding site positioned between the alpha-amino group of Val 1 alpha and the guanidinium group of Arg 141 alpha. Gly-NH2(142 alpha) Hb restores normal oxygen binding properties upon trypsin digestion which removes Gly-NH2 from the COOH terminus of the alpha subunit.