EFFECT OF pH ON THE RATE OF FORMATION AND ON THE EQUILIBRIUM CONCENTRATION OF THE CARBANION INTERMEDIATE By ENRICO GRAZI

The rate of oxidation by ferricyanide of the aldolase-dihydroxyacetone phosphate complex was measured under different conditions. The following conclusions are drawn. 1. In the cleavage of fructose diphosphate, catalysed by native aldolase, the steady-state concentration of the enzyme-dihydroxyacetone phosphate carbanion intermediate represents less than 6% of the total enzyme-substrate intermediates. 2. Fructose diphosphate and dihydroxyacetone phosphate compete for the four catalytic sites on aldolase, the binding offructose diphosphate being about twice as tight. 3. The equilibrium concentration of the carbanion intermediate formed by reaction of carboxypeptidase-treated aldolase with dihydroxyacetone phosphate is independent ofpH between 5.0 and 9.0. The rates offormation ofthe carbanion intermediate and ofthe reverse reaction are, however, concomitantly increased by increasing pH between 5.0 and 6.5.