Journal
CHEMBIOCHEM
Volume 4, Issue 8, Pages 721-726Publisher
WILEY-V C H VERLAG GMBH
DOI: 10.1002/cbic.200200475
Keywords
asymmetric synthesis; directed evolution; enzyme catalysis; hydroxy ketones; protein engineering
Ask authors/readers for more resources
Alteration of the substrate specificity of thiamin diphosphate (ThDP)-dependent benzoylformate decarboxylase (BFD) by error-prone PCR is described. Two mutant enzymes, L476O and M365L-L4615, were identified that accept ortho-substituted benzaldehyde derivatives as donor substrates, which leads to the formation of 2-hydroxy ketones. Both variants, L476O and M365L-L4615 selectively catalyze the formation of enantiopure (S)-2-hydroxy-1-(2-methylphenyl)propan-1-one with excellent yields, a reaction which is only poorly catalyzed by the wild-type enzyme. Different ortho-substituted benzaldehyde derivatives, such as 2-chloro-, 2-methoxy-, or 2-bromobenzaldehyde are accepted as donor substrates by both BFD variants as well and conversion with acetaldehyde resulted in the corresponding (S)-2-hydroxy-1-phenyl-propan-1-one derivatives. As deduced from modeling studies based on the 3D structure of wild type BFD, reduction of the side chain size at position L461 probably results in an enlarged substrate biding site and facilitates the initial binding of ortho substituted benzaldehyde derivatives to the cofactor ThDP.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available