Pyruvate carboxylase from chicken liver. Steady state kinetic studies indicate a "two-site" ping-pong mechanism.

On the basis of initial velocity and product inhibition studies a nonclassical Ping Pong Bi Bi Uni Uni mechanism has been proposed for pyruvate carboxylase from chicken liver. The nonclassical feature of this mechanism is the proposal that each active site on the enzyme is composed of two separate and functionally distinct catalytic sites, i.e., a separate catalytic site exists for the reactants of each partial reaction. The two catalytic sites are presumably linked by the biotinyl residue which functions as the carboxyl carrier. The Bi Bi partial reaction, in which MgATP2-, MgADP-, HCOs-, and phosphate are the substrates, is proposed to utilize a rapid equilibrium random Bi Bi mechanism which includes the formation of two abortive complexes, E-HCOs-Pi and E-HCO,-MgADP-. A rate equation has been developed for the proposed mechanism by employing a combination of rapid equilibrium and steady state methodology. The proposed mechanism is directly analogous to the modsed Ping-Pong Bi Bi mechanism previously described for the biotin-enzyme methylmalonyl-CoA transcarboxylase (D. B. Northrop (1969) J. Biol. Chem., 244, 5808).