Rational Design of Hybrid Natural Products by Utilizing the Promiscuity of an Amide Synthetase.

WS9326A and annimycin are produced by Streptomyces asterosporus DSM 41452. WS9326A is a nonribosomal peptide synthetase-(NRPS-) derived depsipeptide containing a cinnamoyl moiety, while annimycin is a linear polyketide bearing a 2-amino-3-hydroxycyclopent-2-enone (C5N) group. Both gene clusters have been sequenced and annotated. In this study, we show that the amide synthetase Ann1, responsible for attaching the C5N unit during annimycin biosynthesis, has the ability to catalyze fortuitous side reactions to polyenoic acids in addition to its main reaction. Novel compounds were generated by feeding experiments and in vitro studies. We also rationally designed a hybrid natural product consisting of the cinnamoyl moiety of WS9326A and the C5N moiety of annimycin by creating a mutant of S. asterosporus that retains genes encoding biosynthesis of the C5N unit of annimycin and the cinnamoyl group of WS9326A. The promiscuity of Ann1 also proved useful for trapping compounds that arise from acyl-ACP intermediates, which occur in the biosynthesis of the cinnamoyl moiety of WS9326A, by hydrolysis. In this pathway, we postulate that sas27 and sas28 genes are involved in the biosynthesis of the cinnamoyl moiety in WS9326A.