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

Initial rate studies were carried out using solubilized bovine brain hexokinase. Both ATP 4and 0i, y-5'-adenylyl methylene diphosphonate are competitive inhibitors of MgATP 2and mixed inhibitors of the sugar substrate, while N-acetylglucosamine, a competitive inhibitor of fructose, is a mixed inhibitor of MgATP 2. 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 ' C-glucose6-P, ADP, and elevated levels of Mg .