ABSTRACT
An attempt was made to gain insight into the mechanism of orthophosphate attenuation of glucose-6-P inhibition of bovine brain hexokinase I (ADP:D-hexose 6-phosphotransferase, EC 2.7.1.1) from experiments of ligand binding and initial rate kinetics. Studies of glucose-6-P and phosphate binding to hexokinase reveal one binding site per hexokinase molecule. A model is presented which is consistent with the binding and kinetic data currently available on the alleviation of glucose-6-P inhibition of brain hexokinase by phosphate. The model implies that hexokinase may exist in equilibrium either as a free or phosphate-associated enzyme. The kinetic parameters of the two enzyme forms are similar except in their ability to bind glucose-6-P. It is suggested that the dissociation constant for glucose-6-P is relatively very high for hexokinase to which phosphate is bound. Phosphate appears to bind at an allosteric site on the enzyme, whereas glucose-6-P is associated either at the active site or at an allosteric site which overlaps the catalytic site.
