A 5000-fold increase in the specificity of a bacterial phosphotriesterase for malathion through combinatorial active site mutagenesis.

ABSTRACT: Phosphotriesterases (PTEs) have been isolated from a range of bacterial species, including Agrobcaterium radiobacter (PTEAr), and are efficient enzymes with broad substrate ranges. The turnover rate of PTEAr for the common organophosphorous insecticide malathion is lower than expected based on its physical properties; principally the pka of its leaving group. In this study, we rationalise the turnover rate of PTEAr for malathion using computational docking of the substrate into a high resolution crystal structure of the enzyme, suggesting that malathion is too large for the PTEAr binding pocket. Protein engineering through combinatorial active site saturation testing (CASTing) was then used to increase the rate of malathion turnover. Variants from a CASTing library in which Ser308 and Tyr3

SUBMITTER: Naqvi T 

PROVIDER: S-EPMC3983157 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature