Cloning And Functional Analysis Of The Benzonaphthyridine Alkaloid Biosynthetic Gene Cluster In Streptomyces Albogriseolus

Streptomyces albogriseolus MGR072 was isolated from the national mangrove reserve in the Fujian Province of China. Compound 1 produced by MGR072 is a novel benzo naphthyridine, its systematic name is 1-N- methyl-3-methylamino-[N-butanoic acid-3?-(9?-methyl-8?-propen-7?-one) -amide]-benzo[f]naphthyridine-2-one. The benzonaphthyridines are structurally unique and bioactive diverse alkaloids. Such compounds in large amount for medicinal research are mainly acquired by synthetic method and reproducing, research on biosynthesis is nothing much.We found the biosynthesis related gene cluster in the genome by bioinformatics analysis of the genome sequence. We cloned the gene cluster, constructed vector and made a long fragment deletion mutant, after analysis of fermentation product, we found the mutant can't produce compound 1; knocked single genes bnr1, bnr2 and bnr3 of the cluster by PCR targeting, acquired mutant strain LMO09B611, LMO09B612 and LMO09B613; cloned single bnr gene which had been knocked to vector p2170, and complemented single genes to the MGR072 genome.After fermentation of wild type, single gene mutant and complementary strain, we found mutant of bnr1 and bnr2 can't produce compound 1, mutant of bnr3 produce little compound 1, which is probably because other Bnr3 activity is exist.This study tried to resolve the biosynthesis pathway of benzonaphthyridine. Through the genetic manipulation of the biosynthetic genes to lay a foundation of benzonaphthyridines biosynthesis and offer opportunity for searching bioactive novel compounds.