Characterization of site-specific mutations in a short-chain-length/medium-chain-length polyhydroxyalkanoate synthase: in vivo and in vitro studies of enzymatic activity and substrate specificity.

ABSTRACT: Saturation point mutagenesis was carried out at position 479 in the polyhydroxyalkanoate (PHA) synthase from Chromobacterium sp. strain USM2 (PhaC(Cs)) with specificities for short-chain-length (SCL) [(R)-3-hydroxybutyrate (3HB) and (R)-3-hydroxyvalerate (3HV)] and medium-chain-length (MCL) [(R)-3-hydroxyhexanoate (3HHx)] monomers in an effort to enhance the specificity of the enzyme for 3HHx. A maximum 4-fold increase in 3HHx incorporation and a 1.6-fold increase in PHA biosynthesis, more than the wild-type synthase, was achieved using selected mutant synthases. These increases were subsequently correlated with improved synthase activity and increased preference of PhaC(Cs) for 3HHx monomers. We found that substitutions with uncharged residues were beneficial, as they resulted in enhanced

SUBMITTER: Chuah JA 

PROVIDER: S-EPMC3675958 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature