title: RegB kinase activity is controlled in part by monitoring the ratio of oxidized to reduced ubiquinones in the ubiquinone pool.
abstract: RegB is a membrane-spanning sensor kinase responsible for redox regulation of a wide variety of metabolic processes in numerous proteobacterial species. Here we show that full-length RegB purified from Escherichia coli membranes contains bound ubiquinone. Four conserved residues in the membrane-spanning domain of RegB are shown to have important roles in ubiquinone binding in vitro and redox sensing in vivo. Isothermal titration calorimetry measurements, coupled with kinase assays under oxidizing and reducing conditions, indicate that RegB weakly binds both oxidized ubiquinone and reduced ubiquinone (ubiquinol) with nearly equal affinity and that oxidized ubiquinone inhibits kinase activity without promoting a redox reaction. We propose a model in which ubiquinone/ubiquinol bound to RegB readily equilibrates with ubiquinones/ubiquinols in the membrane, allowing the kinase activity to be tuned by the redox state of the ubiquinone pool. This noncatalytic role of ubiquinone in controlling RegB activity is distinct from that of other known ubiquinone-binding proteins, which use ubiquinone as an electron donor or acceptor.
authors.affiliation: Department of Molecular and Cellular Biochemistry, Indiana University, Bloomington, Indiana, USA.
authors.forename: Jiang
authors.initials: J
authors.lastname: Wu
authors.forename: Carl E
authors.initials: CE
authors.lastname: Bauer
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D008954:Models, Biological; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D011485:Protein Binding; D011494:Protein Kinases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D014451:Ubiquinone / Q000031:analogs & derivatives* / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; C091081:RegB protein, Rhodobacter; D014451:Ubiquinone; D011494:Protein Kinases; C003741:ubiquinol