Kinetics of Activation of Acetyl-coA RELATIONSHIP TO THE RATE OF POLYMERIZATION Carboxylase by Citrate OF THE ENZYME*

The kinetics of citrate-induced activation and polym- erization (into filaments) of the 450,000-dalton protomeric form of acetyl-coA carboxylase were com-pared to assess the concertedness of the two processes. Rapid-quench techniques were employed to measure the time course of activation by citrate of the carbox- ylase-catalyzed reaction. When enzyme was preincubated with citrate prior to initiating the steady state turnover reaction with acetyl-coA in the rapid-quench device, the observed rate of carboxylation of acetyl- CoA was apparently linear from the moment of mixing. However, when enzyme was mixed with citrate to initiate the reaction, a lag (tLh = 0.7 S ) occurred in the approach to steady state carboxylation rate. This lag was independent of enzyme concentration over a 230- fold range and was marginally dependent upon citrate concentration. Over the same range of enzyme concentration, polymerization of carboxylase protomers, as determined by right angle light scattering, was enzyme concentra-tion-dependent in a manner predicted by a single pro- tomer activation step, followed by a rate-limiting dimerization of active protomer and subsequent polym- erization. Based on these results, it is concluded that activation of catalysis and the polymerization of car- boxylase protomers are not concerted. Furthermore, activation of carboxylation leading to the formation of an active protomer was faster than polymerization under all conditions, and therefore precedes polymerization.