Early Cephamycin Biosynthetic Genes Are Expressed from a Polycistronic Transcript in Streptomyces clavuligerus

A polycistronic transcript that is initiated at thelat promoter has been implicated in the expression of the genes involved in early steps of cephamycin C biosynthesis inStreptomyces clavuligerus. pcbC is also expressed as a monocistronic transcript from its own promoter. However, an alternative interpretation involving expression via three separate yet interdependent transcripts has also been proposed. To distinguish between these possibilities, mutants lacking thelat promoter and containing a transcription terminator within thelat gene (Δlat ::tsr /term mutants) were created. This mutation eliminated the production of lysine-ɛ-aminotransferase (thelat gene product) but also affected the expression of downstream genes, indicating an operon arrangement. Production of δ-(l -α-aminoadipyl)-l -cysteinyl-d -valine synthetase (ACVS) (thepcbAB gene product) was eliminated in Δlat ::tsr /term mutants, while production of isopenicillin N synthase (IPNS) (thepcbC gene product) was greatly reduced. The provision of α-aminoadipate to the Δlat ::tsr /term mutants, either via exogenous feeding or vialat gene complementation, did not restore production of ACVS or IPNS. Analysis of RNA isolated from the Δlat ::tsr /term mutants confirmed that the polycistronic transcript was absent but also indicated that monocistronicpcbC transcript levels were greatly decreased. In contrast, Δlat mutants created by in-frame internal deletion oflat maintained the polycistronic transcript and allowed production of wild-type levels of both ACVS and IPNS.