Thioesterases and the premature termination of polyketide chain elongation in rifamycin B biosynthesis by Amycolatopsis mediterranei S699

Abstract

The role of two thioesterase genes in the premature release of polyketide synthase intermediates during rifamycin biosynthesis in the Amycolatopsis mediterranei S699 strain was investigated. Creation of an in-frame deletion in the rifR gene led to a 30 ~ 60% decrease in the production of both rifamycin B by the S699 strain or a series of tetra- to decaketide shunt products of polyketide chain assembly by the rifF strain. Since a similar percentage decrease was seen in both genetic backgrounds, we conclude that the RifR thioesterase 2 is not involved in premature release of the carbon chain assembly intermediates. Similarly, fusion of the Saccharopolyspora erythraea DEBS3 thioesterase 1 domain to the C-terminus of the RifE PKS subunit did not result in a noticeable increase in the amount of the undecaketide intermediate formed nor in the amounts of the tetra- to decaketide shunt products. Hence, premature release of the carbon chain assembly intermediates is an unusual property of the Rif PKS itself.