Identification of novel methylbenzene-containing polyketides produced by an actinomycete with rifampicin-induced rpoB gene mutation

Actinomycetes are a group of high G+C, Gram-positive bacteria renowned for their capability of producing a wide range of bioactive secondary metabolites. Many compounds with potent bioactivities such as antitumor and antibacterial activities have been isolated from actinomycetes. However, in recent years, conventional screening for novel bioactive compounds often results in the rediscovery of known compounds. On the contrary, genome sequencing disclosed that each actinomycete has more than 20 biosynthetic gene clusters predicted to encode different structural classes of secondary metabolites. Intriguingly, most of the predicted gene clusters are classified as cryptic or orphan, because the metabolites produced by these cryptic gene clusters are unidentified. To explore these potential novel compounds, we implemented a cryptic gene activation strategy by screening of mutants conferring rifampicin resistance. Induction of rifampicin resistance in 11 actinomycete strains generated 164 rifampicin resistant-mutants (rif mutants). Comparative metabolic screening of these rif mutants with their wild type strains indicated that a mutant (TW-R50-13) overproduced 2 unidentified metabolites. Isolation and structural elucidation of these metabolites revealed that they are novel compounds with a methylbenzene moiety, the biosynthesis of which, to the best of our knowledge, has not been reported. Feeding experiment using ¹³C-labeled precursors demonstrated that the methylbenzene moiety is most likely a product by the action of polyketide synthase. Further investigations on the biosynthesis of these novel compounds are still in progress.