title: Molecular and biochemical characterization of a novel two-component signal transduction system, amrA- amkA, involved in rifamycin SV production in Amycolatopsis mediterranei U32.
abstract: Two-component and phosphorelay signal transduction systems are the major means by which bacteria recognize and respond to a variety of environmental stimuli. Although several model systems, including sporulation in Bacillus subtilis and chemotaxis in Escherichia coli, have been extensively studied, the two-component signal transduction systems in industrially important actinomycetes are not well studied. We report the molecular and biochemical characterization of a novel two-component signal system, amrA-amkA,from the rifamycin-SV-producing Amycolatopsis mediterranei U32. The deduced sequences of amkAand amrA contain all the structural features that are highly conserved in the typical bacterial histidine kinases and response regulators, respectively. BLAST analyses showed that AmrA and AmkA displayed high similarities to AfsQ1/AfsQ2 of Streptomyces coelicolor and MtrA/MtrB of Mycobacterium tuberculosis. The amrAand amkA genes were over-expressed and the gene products were purified from E. coli. Biochemical studies showed that AmkA is able to autophosphorylate, supporting its functional assignment as a histidine kinase. That AmrA functions as the cognate response regulator for histidine kinase AmkA was demonstrated by in vitro phosphotransfer from [gamma-(32)P]ATP-labeled AmkA to AmrA. Rifamycin SV production was also decreased by 10-20% in amrAor amkA gene disruption mutants under the tested condition. Although the detailed regulatory mechanism is still unknown, this is the first report regarding the involvement of two-component signal systems in rifamycin biosynthesis in the genus Amycolatopsis.
authors.affiliation: Laboratory of Molecular Microbiology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 300 Fenglin Road, Shanghai, 200032, P R China.
authors.forename: Weiwu
authors.initials: W
authors.lastname: Wang
authors.forename: Weiwen
authors.initials: W
authors.lastname: Zhang
authors.forename: Hui
authors.initials: H
authors.lastname: Chen
authors.forename: Juishen
authors.initials: J
authors.lastname: Chiao
authors.forename: Guoping
authors.initials: G
authors.lastname: Zhao
authors.forename: Weihong
authors.initials: W
authors.lastname: Jiang
mesh_terms: D000192:Actinomycetales / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D000071677:Histidine Kinase; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D012294:Rifamycins / Q000096:biosynthesis*; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D012294:Rifamycins; C023808:rifamycin SV; D011494:Protein Kinases; D000071677:Histidine Kinase