Structural comparison of lactate dehydrogenase homologs differing in sensitivity to hydrostatic pressure

Abstract

The muscle-type (M 4) lactate dehydrogenases ( l-lactate:NAD + oxidoreductase EC 1.1.1.27) of two teleost fishes, Sebastolobus alascanus and Sebastolobus altivelis, differ in the susceptibility of ligand binding to perturbation by moderate hydrostatic pressures. The enzyme homologs were purufied by affinity chromatography. The amino-acid compositions of these enzymes are virtually identical. The proteins were digested with trypsin and the peptide mixtures mapped using reverse-phase HPLC. Although there was variation in elution times of some peaks, the amino-acid compositions of the fractions from the two profiles were highly similar. Only one clear difference in amino-acid composition was found and this peptide was sequenced using the manual dansyl-Edman method. The enzyme of S. alascanus, which is susceptible to pressure-perturbation, had a histidine at position 115; the S. altivelis enzyme had an asparagine. Ionization of histidine is affected by pressure and may be involved in the differences between the two lactate dehydrogenase homologs. There is no covalently bound phosphate associated with either enzyme, and thus phosphorylation cannot account for the differences between the enzyme homologs. Acquisition of pressure-tolerance appears to involve only minor changes in primary structure.