Purification and characterization of the recombinant Na+-translocating NADH : quinone oxidoreductase from vibrio cholerae

摘要: The nqr operon from Vibrio cholerae, encoding the entire six-subunit, membrane-associated, Na+-translocating NADH:quinone oxidoreductase (Na+-NQR), was cloned under regulation of PBAD promoter. enzyme successfully expressed in V. cholerae. To facilitate molecular genetics studies this sodium-pumping enzyme, a host strain cholerae constructed which genomic copy deleted. By using vector containing six-histidine tag on carboxy terminus NqrF subunit, last subunit operon, recombinant readily purified by affinity chromatography highly active form detergent-solubilized membranes has high specific activity presence sodium. NADH consumption assessed at turnover number 720 electrons per second. When dodecyl maltoside (DM), isolated contains approximately one bound ubiquinone, whereas if detergent LDAO is used instead, t...