Functional Characterization of a Dehydratase Domain from the Pikromycin Polyketide Synthase.

ABSTRACT: Metabolic engineering of polyketide synthase (PKS) pathways represents a promising approach to natural products discovery. The dehydratase (DH) domains of PKSs, which generate an α,β-unsaturated bond through a dehydration reaction, have been poorly studied compared with other domains, likely because of the simple nature of the chemical reaction they catalyze and the lack of a convenient assay to measure substrate turnover. Herein we report the first steady-state kinetic analysis of a PKS DH domain employing LC-MS/MS analysis for product quantitation. PikDH2 was selected as a model DH domain. Its substrate specificity and mechanism were interrogated with a systematic series of synthetic triketide substrates containing a nonhydrolyzable thioether linkage as well as by site-directed mutagenes

SUBMITTER: Li Y 

PROVIDER: S-EPMC4624270 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature