Crystal structure of cytoplasmic acetoacetyl-CoA thiolase from Saccharomyces cerevisiae

Abstract

Thiolases are vital enzymes which participate in both degradative and biosynthetic pathways. Biosynthetic thiolases catalyze carbon-carbon bond formation by a Claisen condensation reaction. The cytoplasmic acetoacetyl-CoA thiolase from Saccharomyces cerevisiae, ERG10, catalyses carbon-carbon bond formation in the mevalonate pathway. The structure of a S. cerevisiae biosynthetic thiolase has not previously been reported. Here, crystal structures of apo ERG10 and its Cys91Ala variant were solved at resolutions of 2.2 and 1.95 , respectively. The structure determined shows that ERG10 shares the characteristic thiolase superfamily fold, with a similar active-site architecture to those of type II thiolases and a similar binding pocket, apart from Ala159 at the entrance to the pantetheine-binding cavity, which appears to be a determinant of the poor binding ability of the substrate. Moreover, comparative binding-pocket analysis of molecule B in the asymmetric unit of the apo structure with that of the CoA-bound complex of human mitochondrial acetoacetyl-CoA thiolase indicates the canonical binding mode of CoA. Furthermore, the steric hindrance revealed in a structural comparison of molecule A with the CoA-bound form raise the possibility of conformational changes that are associated with substrate binding.Structural comparison of the apo structure of S. cerevisiae thiolase with the structure of the CoA-bound complex of human mitochondrial acetoacetyl-CoA thiolase raises the alternate hypothesis that its identical ordered conformation is induced by substrate binding.