title: Identification of a cluster-situated activator of oxytetracycline biosynthesis and manipulation of its expression for improved oxytetracycline production in Streptomyces rimosus.
abstract: BACKGROUND
  Oxytetracycline (OTC) is a broad-spectrum antibiotic commercially produced by Streptomyces rimosus. Despite its importance, little is known about the regulation of OTC biosynthesis, which hampered any effort to improve OTC production via engineering regulatory genes.
  
  
  RESULTS
  A gene encoding a Streptomyces antibiotic regulatory protein (SARP) was discovered immediately adjacent to the otrB gene of oxy cluster in S. rimosus and designated otcR. Deletion and complementation of otcR abolished or restored OTC production, respectively, indicating that otcR encodes an essential activator of OTC biosynthesis. Then, the predicted consensus SARP-binding sequences were extracted from the promoter regions of oxy cluster. Transcriptional analysis in a heterologous GFP reporter system demonstrated that OtcR directly activated the transcription of five oxy promoters in E. coli, further mutational analysis of a SARP-binding sequence of oxyI promoter proved that OtcR directly interacted with the consensus repeats. Therefore, otcR was chosen as an engineering target, OTC production was significantly increased by overexpression of otcR as tandem copies each under the control of strong SF14 promoter.
  
  
  CONCLUSIONS
  A SARP activator, OtcR, was identified in oxy cluster of S. rimosus; it was shown to directly activate five promoters from oxy cluster. Overexpression of otcR at an appropriate level dramatically increased OTC production by 6.49 times compared to the parental strain, thus demonstrating the great potential of manipulating OtcR to improve the yield of OTC production.
authors.affiliation: Department of Environmental and Biological Engineering, School of Chemical and Environmental Engineering, China University of Mining and Technology (Beijing), D11 Xueyuan Road, Haidian District, Beijing, 100083, People's Republic of China. yinsl@student.cumtb.edu.cn.
authors.forename: Shouliang
authors.initials: S
authors.lastname: Yin
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, 1 Beichen West Road, Chaoyang District, Beijing, 100101, People's Republic of China. wangws@im.ac.cn.
authors.forename: Weishan
authors.initials: W
authors.lastname: Wang
authors.affiliation: Shengxue Dacheng Pharmaceutical Co., Ltd, 50 Shengxue Road, Shijiazhuang, 051430, Hebei, People's Republic of China. wangxuefeng1982@163.com.
authors.forename: Xuefeng
authors.initials: X
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, 1 Beichen West Road, Chaoyang District, Beijing, 100101, People's Republic of China. zhuyx@im.ac.cn.
authors.forename: Yaxin
authors.initials: Y
authors.lastname: Zhu
authors.affiliation: Shengxue Dacheng Pharmaceutical Co., Ltd, 50 Shengxue Road, Shijiazhuang, 051430, Hebei, People's Republic of China. nj_jxl@126.com.
authors.forename: Xiaole
authors.initials: X
authors.lastname: Jia
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, 1 Beichen West Road, Chaoyang District, Beijing, 100101, People's Republic of China. lishanshan8427@gmail.com.
authors.forename: Shanshan
authors.initials: S
authors.lastname: Li
authors.affiliation: Shengxue Dacheng Pharmaceutical Co., Ltd, 50 Shengxue Road, Shijiazhuang, 051430, Hebei, People's Republic of China. bcd163@163.com.
authors.forename: Fang
authors.initials: F
authors.lastname: Yuan
authors.affiliation: Department of Environmental and Biological Engineering, School of Chemical and Environmental Engineering, China University of Mining and Technology (Beijing), D11 Xueyuan Road, Haidian District, Beijing, 100083, People's Republic of China. zhangyuxiu@cumtb.edu.cn.
authors.forename: Yuxiu
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, 1 Beichen West Road, Chaoyang District, Beijing, 100101, People's Republic of China. yangkq@im.ac.cn.
authors.forename: Keqian
authors.initials: K
authors.lastname: Yang
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics; D009154:Mutation; D010118:Oxytetracycline / Q000096:biosynthesis*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D017386:Sequence Homology, Amino Acid; D064411:Streptomyces rimosus / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D001426:Bacterial Proteins; D010118:Oxytetracycline