Essential carboxyl groups in yeast hexokinase

Although a great deal of kinetic data are available on yeast hexokinase and several schemes have been proposed, the mechanism of its action is far from being understood [ 1,2] . It is generally agreed, however, that the transphosphorylation step requires the formation of a ternary complex between the enzyme and its substrates, ATP-Mg and D-glucose. Although the pH dependence of the enzyme activity suggests that at least one ionisable group, such as the imidazole of histidine or the y-carboxyl group of aspartic or glutamic acids, takes part in the catalytic process [3, 41, the nature of the amino acid residues involved in enzymatic catalysis or substrate binding remains to be determined. According to the results of Grouselle et al. [5], as well as our own unpublished results, the histidine residues do not appear to be associated either with the catalytic step or with the enzyme-substrate interactions. Thus, the inactivation observed when the enzyme is carboethoxylated is caused by a conformational change in the enzyme protein rather than by the modification of an essential histidine residue. It seemed, therefore, important to investigate the effect of specific carboxyl group reagents upon the enzyme activity. Water soluble carbodiimides have been shown to be good reagents for this purpose [6]. This paper reports some preliminary results obtained by chemical modification of essential carboxyl groups in yeast hexokinase.