Reference - PMID:16216079 - Catalytic mechanism of fungal homoserine transacetylase.
Reference summary
- PubMed ID
- PMID:16216079
- Title
- Catalytic mechanism of fungal homoserine transacetylase.
- Authors
- Nazi I, Wright GD
- Citation
- Biochemistry 2005 Oct 18;44(41):13560-6
- Publication year
- 2005
- Abstract
- Homoserine transacetylase is a required catalyst in the biochemical pathway that metabolizes Asp to Met in fungi. The enzyme from the yeast Schizosaccharomyces pombe activates the hydroxyl group of L-homoserine by acetylation from acetyl coenzyme A. This enzyme is unique to fungi and some bacteria and presents an important new target for drug discovery. Steady-state kinetic parameters provide evidence that this enzyme follows a ping-pong mechanism. Proton inventory was consistent with a single-proton transfer, and pH studies suggested the participation of at least one residue with a pKa value of 6.4-6.6, possibly a His or Asp/Glu in catalysis. Protein sequence alignments indicate that this enzyme belongs to the alpha/beta-hydrolase fold superfamily of enzymes, indicating the involvement of an active-site nucleophile and possibly a canonical catalytic triad. We constructed site-specific mutants and identified Ser163, Asp403, and His432 as the likely active-site residues of a catalytic triad based on steady-state kinetics and genetic complementation of a yeast null mutant. Moreover, unlike the wild-type enzyme, inactive site mutants were not capable of producing an acetyl-enzyme intermediate. Homoserine transacetylase therefore catalyzes the acetylation of L-homoserine via a covalent acyl-enzyme intermediate through an active-site Ser. These results form the basis of future exploitation of this enzyme as an antimicrobial target.
