Hysteretic behaviour of citrate synthase. The reaction mechanism and the exclusion of synthase being a hysteretic enzyme.

The non-Michaelis-Menten kinetics, burst and steady-state periods, expressed by citrate synthase in the presence of citryl-CoA, were investigated by labelling experiments with trace amounts of [14C]acetyl-CoA. The results indicate that citrate becomes labelled in the reaction of liberated acetyl-CoA with the binary synthase.oxaloacetate complex that is transiently generated in the lyase reaction of citryl-CoA. Mediated by the hydrolase function of synthase, the counteracting citryl-CoA lyase and ligase reactions operate towards a transient flow equilibrium. This precedes the thermodynamic equilibrium and is established during the burst period; it is maintained under steady-state conditions and corresponds to the formation of transiently nonproductive synthase. The rates of both synthase partial reactions, therefore, are likewise affected. Oxaloacetate in the presence of acetyl-CoA competitively inhibits the hydrolysis of citryl-CoA and vice versa. In the synthase dependence of the burst periods and during the time dependence of the steady-state periods, nonproportionally more of physiological substrates participate in citrate formation. The nonproportional increase is a consequence of the continuously changing conditions to establish or to maintain the flow equilibrium, respectively, during the reaction progress. Third rate periods after the steady state result if the equilibrium conditions cannot be satisfied. High concentrations of oxaloacetate inhibit the expression of non-Michaelis-Menten kinetics by formation of nonproductive synthase.oxaloacetate complex. The supply of acetyl-CoA is then sufficient and the formation of the flow equilibrium prevented. The implication of the results with structural work is discussed.