Mutational analysis of the Paracoccus denitrificans c-type cytochrome biosynthetic genes ccmABCDG: disruption of ccmC has distinct effects suggesting a role for CcmC independent of CcmAB.
Each of the Paracoccus denitrificans genes in the c-type cytochrome biogenesis gene cluster ccmABCDG, plus the two flanking genes ORF117 and hisH, were individually disrupted by omega insertion. Resultant phenotypes were restored to the wild-type by complementation from a set of plasmids. All of the ccm genes, but neither ORF117 nor hisH, were required for c-type cytochrome biogenesis; only ccmG was also implicated in the biosynthesis of cytochrome aa3. Disruption of ccmC or ccmG resulted in failure to grow on rich media, but disruption of ccmA, ccmB or ccmD did not. The ccmC mutant, but not the ccmA, ccmB or ccmD mutants, also exhibited the increased sensitivity to growth inhibition by oxidized thiol compounds previously observed for the ccmG mutant. In contrast to the ccmG mutant, however, growth of the ccmC mutant on rich media could not be restored by DTT. Siderophore biosynthesis and/or secretion by P. denitrificans was also attenuated by disruption of ccmC and ccmG but not of ccmA, ccmB or ccmD. These results indicate that CcmC can function independently of CcmA, CcmB and CcmD despite other evidence that these gene products form an ATP-binding cassette (ABC)-type-transporter with the subunit composition (CcmA)2-CcmB-CcmC or (CcmA)2-CcmB-CcmC-CcmD, and also suggest a possible link between the functions of CcmC and CcmG.
Page
M Dudley
MD
Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK, and Oxford Centre for Molecular Sciences, New Chemistry Building, South Parks Road, Oxford, OX1 3QT, UK1.
Ferguson
Stuart J
SJ
Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK, and Oxford Centre for Molecular Sciences, New Chemistry Building, South Parks Road, Oxford, OX1 3QT, UK1.
D018528:ATP-Binding Cassette Transporters; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000502:physiology*; D001426:Bacterial Proteins; D001483:Base Sequence; D003470:Culture Media / Q000737:chemistry; D003574:Cytochrome c Group / Q000096:biosynthesis / Q000235:genetics*; D004220:Disulfides / Q000494:pharmacology; D003576:Electron Transport Complex IV / Q000096:biosynthesis; D005798:Genes, Bacterial / Q000235:genetics*; D005816:Genetic Complementation Test; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016366:Open Reading Frames / Q000235:genetics; D010088:Oxidoreductases / Q000235:genetics; D010231:Paracoccus denitrificans / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D033903:Periplasmic Proteins; D015183:Restriction Mapping; D017262:Siderophores / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D000637:Transaminases / Q000235:genetics
D018528:ATP-Binding Cassette Transporters; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; C118373:CcmA protein, bacteria; C116246:CcmC protein, bacteria; D003470:Culture Media; D003574:Cytochrome c Group; D004220:Disulfides; D008565:Membrane Proteins; D033903:Periplasmic Proteins; D017262:Siderophores; D010088:Oxidoreductases; C077913:periplasmic protein disulfide oxidoreductase; D003576:Electron Transport Complex IV; D000637:Transaminases; C053430:histidinol-phosphate aminotransferase