Pyruvate carboxylase (PC) catalyzes the biotin-dependent carboxylation of pyruvate to produce oxaloacetate. PC has crucial roles in gluconeogenesis, lipogenesis, glyceroneogenesis, and insulin secretion in mammals. PC deficiencies are linked to lactic acidemia, hypoglycemia, and other diseases. Four single-site mutations, V145A, R451C, A610T, M743I, are associated with these diseases.

PC is a multi-domain enzyme of about 130 kD in eukaryotes and most bacteria. It contains the biotin carboxylase (BC), carboxyltransferase (CT), and biotin-carboxyl carrier protein (BCCP) domains. The BC domain catalyzes the first step of the reaction: the carboxylation of the biotin prosthetic group that is covalently linked to BCCP. In the second step of the reaction, the carboxyltransferase (CT) domain catalyzes the transfer of the carboxyl group from (carboxy)biotin to pyruvate.

PC is only active in the tetrameric form. The catalytic activity is stimulated by acetyl-CoA and inhibited by aspartate.

• The crystal structure of the CT+PT+BCCP domain of human PC has been determined at 2.8A resolution.


• The crystal structure of full-length S. aureus PC has been determined at 2.8A resolution.


• The structures identify a novel domain, the PC tetramerization (PT) domain, that is essential for the tetramerization of this enzyme.


• Mutation of a single residue in the PT domain, F1077A, can disrupt the tetramer and inactivate the enzyme.


• The BCCP domain is swapped between dimers in the tetramer, explaining why only tetramers of the enzyme are active.


• The active sites of the BC and CT domains are separated by about 75 A, suggesting that the entire BCCP domain will need to translocate during the reaction.


• BCCP-biotin is observed in the active site of the CT domain, providing the first molecular insight into how biotin participates in the CT reaction.


• The A610T mutation is likely to block biotin binding in the CT active site, while the M743I mutation is likely to block pyruvate binding.


• The R451C mutation is located in the acetyl-CoA binding site, and de-sensitizes the enzyme towards this activator.


• The V145A mutation is in the BC domain, and is known to destabilize the enzyme.


• There are dramatic conformational differences to the structure of R. etli PC, obtained in complex with the activator ethyl-CoA. The structural differences may be due to the binding of this activator.


• An exo binding site for biotin is observed, located at the PT-CT domain interface. The functional relevance of this site is currently not known.

Publications from this project

• S. Xiang & L. Tong. (2008). Crystal structures of human and Staphylococcus aureus pyruvate carboxylase and molecular insights into the carboxyltransfer reaction. Nature Struct. Mol. Biol., 15, 295-302. Reprint(PDF)

Funding for this project

• NIH R01 DK67238