title: Paracoccus denitrificans CcmG is a periplasmic protein-disulphide oxidoreductase required for c- and aa3-type cytochrome biogenesis; evidence for a reductase role in vivo.
abstract: Cloning and sequencing of the Paracoccus denitrificans ccmG gene indicates that it codes for a periplasmic protein-disulphide oxidoreductase; the presence of the sequence Cys-Pro-Pro-Cys at the CcmG active site suggests that it may act in vivo to reduce disulphide bonds rather than to form them. A CcmG-PhoA fusion confirmed the periplasmic location. Disruption of the ccmG gene resulted in not only the expected phenotype of pleiotropic deficiency in c-type cytochromes, but also loss of spectroscopically detectable cytochrome aa3, cytochrome c oxidase and ascorbate/TMPD oxidase activities; there was also an enhanced sensitivity to growth inhibition by some component of rich media and by oxidized thiol compounds. Dithiothreitol promoted the growth of the ccmG mutant on rich media and substantially restored spectroscopically detectable cytochrome aa3 and cytochrome c oxidase activity, although it did not restore c-type cytochrome biogenesis. Assembly of the disulphide-bridged proteins methanol dehydrogenase and Escherichia coli alkaline phosphatase was unaffected in the ccmG mutant. It is proposed that P. denitrificans CcmG acts in vivo to reduce protein-disulphide bonds in certain protein substrates including c-type cytochrome polypeptides and/or polypeptides involved in c-type cytochrome biogenesis.
authors.affiliation: Department of Biochemistry, University of Oxford, UK. mpage@molbiol.ox.ac.uk
authors.forename: M D
authors.initials: MD
authors.lastname: Page
authors.forename: S J
authors.initials: SJ
authors.lastname: Ferguson
mesh_terms: D000429:Alcohol Oxidoreductases / Q000378:metabolism; D000469:Alkaline Phosphatase / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000502:physiology*; D003001:Cloning, Molecular; D003470:Culture Media; D003574:Cytochrome c Group / Q000096:biosynthesis*; D004229:Dithiothreitol / Q000494:pharmacology; D003576:Electron Transport Complex IV / Q000096:biosynthesis*; D004926:Escherichia coli / Q000201:enzymology; D008969:Molecular Sequence Data; D009154:Mutation; D010088:Oxidoreductases / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D010231:Paracoccus denitrificans / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D033903:Periplasmic Proteins; D011993:Recombinant Fusion Proteins / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013438:Sulfhydryl Compounds / Q000494:pharmacology; D000637:Transaminases / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D003470:Culture Media; D003574:Cytochrome c Group; D033903:Periplasmic Proteins; D011993:Recombinant Fusion Proteins; D013438:Sulfhydryl Compounds; D010088:Oxidoreductases; D000429:Alcohol Oxidoreductases; C019641:alcohol dehydrogenase (acceptor); C077913:periplasmic protein disulfide oxidoreductase; D003576:Electron Transport Complex IV; D000637:Transaminases; C053430:histidinol-phosphate aminotransferase; D000469:Alkaline Phosphatase; D004229:Dithiothreitol