Comparison of Carboxymethylation Patterns of Harbor Seal and Sperm Whale Myoglobins

Abstract

Abstract Harbor seal (Phoca vitulina) and sperm whale (Physeter catodon) myoglobins were carboxymethylated in 0.2 m bromoacetate at pH 6.8. The modification of histidine residues leveled off after approximately 6 and 8 days, respectively, of reaction at 25° in the dark. The NH2-terminal glycine and valine residues, respectively, were modified, as were 1 or 2 lysine residues on the average. The modified seal protein was subjected to digestion with trypsin and the acid-insoluble fraction from that treatment further exposed to thermolysin. The modified sperm whale protein was subjected to thermolysin alone. Peptides containing histidine or a derivative were isolated and identified, and conclusions concerning the patterns of reactivity were tabulated. The results with the sperm whale protein confirm and complete the assignments reported previously (Hugli, T. E., and Gurd, F. R. N. (1970) J. Biol. Chem. 245, 1939–1946). Histidine residue 64 was found to be unreactive, while the peculiar suppression of reactivity of residue 36 relative to the reaction product obtained in the crystalline state was confirmed. The pattern of reactivity of the harbor seal protein was similar. The freely reactive histidine residues were 8, 81, 113, and 116, the unreactive residues were 24, 64, 82, 93, and 97, and those showing various patterns of restriction were 36, 48, 119, and 152.