In vivo investigation of the role of SfmO2 in saframycin A biosynthesis by structural characterization of the analogue saframycin O

Saframycin A（SFM-A）,a tetrahydroisoquinoline antibiotic isolated from Streptomyces lavendulae,shows potent anti-proliferation activities against a variety of tumor cell lines,and shares the core structure with ecteinascidin 743（ET-743）,the anticancer drug for soft-tissue sarcoma.Characterization of the SFM-A biosynthetic gene cluster revealed three nonribosomal peptide synthetase genes and a series of genes encoding oxygenases.To investigate the function of sfmO2 gene,encoding a FAD-dependent monooxygenase/hydroxylase,we constructed the gene replacement mutant（△sfmO2） strain S.lavendulae TL2007 and the corresponding gene complementation mutant strain S.lavendulae TL2008.A novel compound,SFM-O,was isolated from the △sfmO2 replacement mutant strain and its structure was characterized by comparison to the HRMS and NMR spectra of SFM-A.These findings indicated that SfmO2 is responsible for the oxidation of ring A in the biosynthetic pathway of SFM-A,and the new compound SFM-O could be considered as an advanced intermediate in the semisynthesis of ET-743.