Pyruvate Carboxylase - Reaction Mechanism

Reaction Mechanism

The reaction mechanism can be subdivided into two partial reactions (see figure to the right). In the first reaction, ATP is carboxylated to produce carbonic phosphoric anhydride which in turn carboxylates a biotin cofactor that is covalently attached to a lysine residue of the BCCP domain. Carbonic phosphoric anhydride decomposes into carbon dioxide and phosphate prior to attack by the enzyme linked biotin molecule. In most species, this reaction requires acetyl-CoA as an allosteric activator binding to the PT domain. In the second reaction, occurring in the CT domain of an adjacent monomer, carbon dioxide is transferred to the acceptor molecule, pyruvate, to form oxaloacetate. The reaction proceeds via the removal of a proton from pyruvate, by an as yet unidentified active site residue, to generate an enolate intermediate. The enolate intermediate than attacks CO₂ transiently released from the enzyme linked biotin molecule. The resultant oxaloacetate is released. The biotin molecule is protonated by the aforementioned active site residue and released from the active site of the CT domain to be recarboxylated. The major regulator of enzyme activity, acetyl-CoA, stimulates the cleavage of ATP in the first partial reaction and also it has been shown to induce a conformational change in the tetrameric structure of the enzyme.