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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.    Principal Findings  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 His 6 Mur34, 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.

Identification of Mur34 as the Novel Negative Regulator Responsible for the Biosynthesis of Muraymycin in Streptomyces sp. NRRL30471