title: Identification of Mur34 as the novel negative regulator responsible for the biosynthesis of muraymycin in Streptomyces sp. NRRL30471.
abstract: BACKGROUND
  Muraymycin, a potent translocase I (MraY) inhibitor, is produced by Streptomyces sp. NRRL30471. The muraymycin gene cluster (mur) was recently cloned, and bioinformatic analysis of mur34 revealed its encoding product exhibits high homology to a large family of proteins, including KanI and RacI in individual biosynthetic pathway of kanamycin and ribostamycin. However, the precise role of these proteins remains unknown.
  
  
  RESULTS
  Here we report the identification of Mur34 as the novel negative regulator involved in muraymycin biosynthesis. Independent disruption of mur34 on chromosome and cosmid directly resulted in significant improvement of muraymycin production by at least 10 folds, thereof confirming the negative function of Mur34 during muraymycin biosynthesis and realizing the engineered production of muraymycin in heterologous host. Gene expression analysis indicated that the transcription level of the mur genes in mur34 mutant (DM-5) was dramatically enhanced by ca. 30 folds. Electrophoretic mobility shift assay (EMSA) showed that Mur34 specifically bound to the promoter region of mur33. Further experiments showed that a 28-bp region downstream of the transcription start point (TSP) was protected by His6Mur34, and the -10 region is essential for the activity of mur33 promoter.
  
  
  CONCLUSIONS
  Mur34 plays an unambiguously negative role in muraymycin biosynthesis via binding to the upstream of mur33. More importantly, Mur34 represents a novel family of regulators acting in negative manner to regulate the secondary metabolites biosynthesis in bacteria.
authors.affiliation: State Key Laboratory of Microbial Metabolism, and School of Life Sciences & Biotechnology, Shanghai Jiao Tong University, Shanghai, China ; NCPC New Drug Research and Development Co., Ltd, North China Pharmaceutical Group Corporation, Shijiazhuang, China.
authors.forename: Dongmei
authors.initials: D
authors.lastname: Xu
authors.forename: Guang
authors.initials: G
authors.lastname: Liu
authors.forename: Lin
authors.initials: L
authors.lastname: Cheng
authors.forename: Xinhua
authors.initials: X
authors.lastname: Lu
authors.forename: Wenqing
authors.initials: W
authors.lastname: Chen
authors.forename: Zixin
authors.initials: Z
authors.lastname: Deng
mesh_terms: D000900:Anti-Bacterial Agents / Q000096:biosynthesis*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D050557:Catechol 2,3-Dioxygenase / Q000378:metabolism; D019295:Computational Biology; D008969:Molecular Sequence Data; D005810:Multigene Family; D009154:Mutation; D009705:Nucleosides / Q000096:biosynthesis*; D011401:Promoter Regions, Genetic; D013302:Streptomyces / Q000235:genetics / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D009705:Nucleosides; D050557:Catechol 2,3-Dioxygenase