Studies on the kinetic mechanism and allosteric nature of bovine brain hexokinase.

Abstract Initial rate studies were carried out using solubilized bovine brain hexokinase. Both ATP4- and β,γ-5'-adenylyl methylene diphosphonate are competitive inhibitors of MgATP2- and mixed inhibitors of the sugar substrate, while N-acetylglucosamine, a competitive inhibitor of fructose, is a mixed inhibitor of MgATP2-. These results are consistent with a random Bi Bi mechanism for the brain enzyme. Purine nucleotides, with the exception of ATP, bind at the catalytic site as well as at an allosteric site on the enzyme. ATP and pyrimidine nucleotides bind only at the catalytic site of brain hexokinase. All allosteric effectors were found to be inhibitory. These include: ADP, AMP, GTP, GDP, ITP, IDP, IMP, 3',5'-cyclic AMP, and 3-phosphoglycerate. The most potent inhibitor of the enzyme appears to be ADP. The hexokinase reaction was reversed using 14C-glucose-6-P, ADP, and elevated levels of Mg2+.