Equilibrium and Net Reaction Rates in Relation to the Mechanism of Yeast Hexokinase

Abstract

Recent reports on the mode of enzyme and substrate interaction of the yeast hexokinase reaction are consistent with the view that substrates n-glucose and adenosine 5'-triphosphate must reside at the catalytically active site of the enzyme prior to product formation (l-4).Hammes and Kochavi (I, 2) concluded from analysis of initial rate data that the reaction mechanism is ordered and that glucose is the obligatory leading substrate.Subsequently, Fromm and Zewe (3) suggested from initial rate studies that substrates may add to hexokinase in a noncompulsory order, leading to ternary complexes in which all steps in the reaction sequence are in rapid equilibrium relative to interconversion of the ternary complexes.Additional support for this suggestion comes from studies by Zewe, Fromm, and Fabian0 (4) of competitive inhibitors, alternate substrates, and the effect of substrates on the quenching of hexokinase fluorescence.Support for both suggested mechanisms is currently available in the literature from nonkinetic investigations.Trayser and Colowick (5) have shown that glucose is capable of protecting the active site of yeast hexokinase against tryptic digestion in the absence of ATP.Cohn (6) has reported from nuclear magnetic resonance experiments that glucose must be present before hexokinase can produce an enhancement of the relaxation rate of MnADP-.These observations were taken to mean that the mechanism of the hexokinase reaction is ordered with glucose as initial substrate.Kaji, Trayser, and Colowick (7) have observed that highly purified yeast hexokinase exhibits adenosine triphosphatase activity and that this activity could be inhibited by substances that probably bind at the glucose site.Such evidence is consistent with the suggestion that ATP may add to the active site of the enzyme in the absence of the sugar substrate (3,4).