Kinetic studies with liver galactokinase

Abstract

Kinetic measurements of the forward reaction catalyzed by ATP [adenosine triphosphate]-galactose phos-photransferase were carried out with a purified preparation from pig liver. The rate of reaction at pH 7.8 is dependent on the concentration of MgATP2- rather than total ATP or MgCl2 concentrations. The effect of changes in pH on Km (galactose), Km (MgATP2-) and Vmax was studied. Of several possible nucleotide substrates only ATP and deoxyATP were effective. The initial-velocity patterns both in the absence and presence of products were determined. Galactose 1-phosphate is a non-competitive inhibitor when either galactose or MgATP2- was the variable substrate. MgADP- was a non-competitive inhibitor with galactose and a competitive inhibitor with MgATP2- as variable substrate. These results are consistent with an ordered reaction pathway in which galactose combines with an initial enzyme-MgATP2- complex.