Structure of the biotin carboxylase subunit of pyruvate carboxylase from Aquifex aeolicus at 2.2 Å resolution

Pyruvate carboxylase (PC) is distributed in many eukaryotes as well as in some prokaryotes. PC catalyzes the ATP-dependent carboxylation of pyruvate to form oxalacetate. PC has three functional domains, one of which is a biotin carboxylase (BC) domain. The BC subunit of PC from Aquifex aeolicus (PC-β) was crystallized in an orthorhombic form with space group P2₁2₁2, unit-cell parameters a = 92.4, b = 122.1, c = 59.0 Å and one molecule in the asymmetric unit. Diffraction data were collected at 100 K on BL24XU at SPring-8. The crystal structure was determined by the molecular-replacement method and refined against 20.0–2.2 Å resolution data, giving an R factor of 0.199 and a free R factor of 0.236. The crystal structure revealed that PC-β forms a dimeric quaternary structure consisting of two molecules related by crystallographic twofold symmetry. The overall structure of PC-β is similar to other biotin-dependent carboxylases, such as acetyl-CoA carboxylase (ACC). Although some parts of domain B were disordered in ACC, the corresponding parts of PC-β were clearly determined in the crystal structure. From comparison between the active-site structure of ACC with ATP bound and a virtual model of PC-β with ATP bound, it was shown that the backbone torsion angles of Glu203 in PC-β change and some of water molecules in the active site of PC-β are excluded upon ATP binding.