Functional and transcriptional analyses of a fengycin synthetase gene, fenC, from Bacillus subtilis

A 37-kb DNA fragment containing five fengycin synthetase genes, including f enC, fenD, fenE, fenA, and fenB, was cloned and sequenced. Among these gene s,fenC encodes a fengycin synthetase 2,560 amino acids long with an estimat ed molecular mass of 287 kDa. This protein contains two amino acid activati on modules, FenC1 and FenC2, which activate L-glutamic acid and L-ornithine , respectively. Primer extension, using mRNA isolated from the log-phase ce lls, identified a transcription start site located 86 nucleotides upstream from the initiation codon of fenC, implying that a promoter is located upst ream from the start site. Primer extension using total RNA isolated from st ationary-phase cells also identified a transcription start site located 61 nucleotides upstream from the initiation codon of fenC. Gene fusion studies demonstrated that in nHA medium, the cells transcribe the fengycin synthet ase genes at two different stages of cell growth. The promoter is active du ring the log phase, and the activity reaches the highest level during the l ate log phase. The activity decreases sharply but is maintained at a low le vel for approximately 24 h after cells enter the early stationary phase. Th e results of this investigation also suggest that the transcription of fenC is positively regulated during the late log phase. Results presented herei n provide further insight into fengycin synthesis by B. subtilis F29-3.