tailieunhanh - Báo cáo khoa học: DNA-binding characteristics of the regulator SenR in response to phosphorylation by the sensor histidine autokinase SenS from Streptomyces reticuli

The two-component system SenS–SenR fromStreptomyces reticulihas been shown to influence the production of the redox regulator FurS, the mycel-ium-associated enzyme CpeB, which displays heme-dependent catalase and peroxidase activity as well as heme-independent manganese peroxidase activity, and the extracellular heme-binding protein HbpS. | ễFEBS Journal DNA-binding characteristics of the regulator SenR in response to phosphorylation by the sensor histidine autokinase SenS from Streptomyces reticuli Gabriele Bogel Hildgund Schrempf and Dario Ortiz de Orue Lucana FB Biologie Chemie Universitat Osnabruck Germany Keywords DNA binding phosphorylation Streptomyces two-component system SenS-SenR Correspondence D. Ortiz de Orue Lucana Universitat Osnabruck FB Biologie Chemie Angewandte Genetik der Mikroorganismen Barbarastr. 13 49069 Osnabruck Germany Fax 49 541 9692804 Tel 49 541 9693439 E-mail ortiz@ Received 13 March 2007 revised 7 June 2007 accepted 7 June 2007 doi The two-component system SenS-SenR from Streptomyces reticuli has been shown to influence the production of the redox regulator FurS the mycelium-associated enzyme CpeB which displays heme-dependent catalase and peroxidase activity as well as heme-independent manganese peroxidase activity and the extracellular heme-binding protein HbpS. In addition it was suggested to participate in the sensing of redox changes. In this work the tagged cytoplasmic domain of SenS SenSc as well as the full-length differently tagged SenR and corresponding mutant proteins carrying specific amino acid exchanges were purified after heterologous expression in Escherichia coli. In vitro SenSc is autophosphorylated to SenSc P at the histidine residue at position 199 transfers the phosphate group to the aspartic acid residue at position 65 in SenR and acts as a phosphatase for SenR P. Bandshift and footprinting assays in combination with competition and mutational analyses revealed that only unphosphorylated SenR binds to specific sites upstream of the furS-cpeB operon. Further specific sites within the regulatory region common to the oppositely orientated senS and hbpS genes were recognized by SenR. Upon its phosphorylation the DNA-binding affinity of this area was enhanced. These data together with previous in