microbiology, Vol 143, 563-576, Copyright © 1997 by Society for General Microbiology

ARTICLES

The Paracoccus denitrificans ccmA, B and C genes: cloning and sequencing, and analysis of the potential of their products to form a haem or apo- c-type cytochrome transporter

MD Page, DA Pearce, HA Norris and SJ Ferguson
Department of Biochemistry, University of Oxford, UK.

Two c-type cytochrome deficient mutants of Paracoccus denitrificans, HN49 and HN53, were isolated by Tn5 mutagenesis and screening for failure to oxidize dimethylphenylenediamine (the Nadi test). Both were completely deficient in c-type cytochromes. Genomic DNA flanking the site of Tn5 insertion in HN53 was cloned by marked rescue and a 3.1 kb region sequenced. Three of the genes, designated ccmA, ccmB and ccmC, present in this region are proposed to encode the components of a membrane transporter of the ABC-(ATP-binding cassette) superfamily, which is similar to a group of transporters postulated to translocate either haem or apocytochromes c. The Tn5 elements in HN49 and HN53 shown to be inserted in ccmB and ccmA, respectively. Sequence analysis suggested that both CcmB and CcmC have the potential to interact with CcmA and thus that the three gene products probably associate to form a complex with (CcmA)2-CcmB-CcmC stoichiometry; it is also indicated a lack of similarity between CcmB and CcmC and the membrane-integral components of transporters mediating uptake of haem or other iron complexes. Supplementation of growth media with haem did not stimulate c-type cytochrome formation in HN49 or HN53, although it elevated levels of soluble haemoproteins and membrane-bound cytochromes b, suggesting that exogenous haem can traverse both outer and inner membranes of P. denitrificans. HN49 and HN53 accumulated apocytochrome C550 to much lower levels than other c-type cytochrome deficient mutants of P. denitrificans but expression and translocation of an apocytochrome C550-alkaline phosphatase fusion protein and apocytochrome cd1 were unaffected in HN53. The results suggest that the substrate for the putative CcmABC-transporter is probably neither haem nor c-type apocytochromes.

This article has been cited by other articles:

				R. Bouhenni, A. Gehrke, and D. Saffarini Identification of Genes Involved in Cytochrome c Biogenesis in Shewanella oneidensis, Using a Modified mariner Transposon Appl. Envir. Microbiol., August 1, 2005; 71(8): 4935 - 4937. [Abstract] [Full Text] [PDF]

				P. P. Hamel, B. W. Dreyfuss, Z. Xie, S. T. Gabilly, and S. Merchant Essential Histidine and Tryptophan Residues in CcsA, a System II Polytopic Cytochrome c Biogenesis Protein J. Biol. Chem., January 17, 2003; 278(4): 2593 - 2603. [Abstract] [Full Text] [PDF]

				B. W. Dreyfuss, P. P. Hamel, S. S. Nakamoto, and S. Merchant Functional Analysis of a Divergent System II Protein, Ccs1, Involved in c-Type Cytochrome Biogenesis J. Biol. Chem., January 17, 2003; 278(4): 2604 - 2613. [Abstract] [Full Text] [PDF]

				O. Daltrop, J. M. Stevens, C. W. Higham, and S. J. Ferguson The CcmE protein of the c-type cytochrome biogenesis system: Unusual in vitro heme incorporation into apo-CcmE and transfer from holo-CcmE to apocytochrome PNAS, July 23, 2002; 99(15): 9703 - 9708. [Abstract] [Full Text] [PDF]

				V. K. Viswanathan, S. Kurtz, L. L. Pedersen, Y. Abu Kwaik, K. Krcmarik, S. Mody, and N. P. Cianciotto The Cytochrome c Maturation Locus of Legionella pneumophila Promotes Iron Assimilation and Intracellular Infection and Contains a Strain-Specific Insertion Sequence Element Infect. Immun., April 1, 2002; 70(4): 1842 - 1852. [Abstract] [Full Text] [PDF]

				R. L. Cox, C. Patterson, and T. J. Donohue Roles for the Rhodobacter sphaeroides CcmA and CcmG Proteins J. Bacteriol., August 1, 2001; 183(15): 4643 - 4647. [Abstract] [Full Text] [PDF]

				G. M. Cook and R. K. Poole Oxidase and periplasmic cytochrome assembly in Escherichia coli K-12: CydDC and CcmAB are not required for haem-membrane association Microbiology, February 1, 2000; 146(2): 527 - 536. [Abstract] [Full Text]

				M. D. Page and S. J. Ferguson 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 Microbiology, November 1, 1999; 145(11): 3047 - 3057. [Abstract] [Full Text]

				H. Schulz, R. A. Fabianek, E. C. Pellicioli, H. Hennecke, and L. Thony-Meyer Heme transfer to the heme chaperone CcmE during cytochrome c maturation requires the CcmC protein, which may function independently of the ABC-transporter CcmAB PNAS, May 25, 1999; 96(11): 6462 - 6467. [Abstract] [Full Text] [PDF]

				S. C. Baker, S. J. Ferguson, B. Ludwig, M. D. Page, O.-M. H. Richter, and R. J. M. van Spanning Molecular Genetics of the Genus Paracoccus: Metabolically Versatile Bacteria with Bioenergetic Flexibility Microbiol. Mol. Biol. Rev., December 1, 1998; 62(4): 1046 - 1078. [Abstract] [Full Text] [PDF]

				R. A. Fabianek, H. Hennecke, and L. Thöny-Meyer The Active-Site Cysteines of the Periplasmic Thioredoxin-Like Protein CcmG of Escherichia coli Are Important but Not Essential for Cytochrome c Maturation In Vivo J. Bacteriol., April 1, 1998; 180(7): 1947 - 1950. [Abstract] [Full Text]

				D. Pearce, M. Page, H. Norris, E. Tomlinson, and S. Ferguson Identification of the contiguous Paracoccus denitrificans ccmF and ccmH genes: disruption of ccmF, encoding a putative transporter, results in formation of an unstable apocytochrome c and deficiency in siderophore production Microbiology, February 1, 1998; 144(2): 467 - 477. [Abstract]

				Q. Ren and L. Thony-Meyer Physical Interaction of CcmC with Heme and the Heme Chaperone CcmE during Cytochrome c Maturation J. Biol. Chem., August 24, 2001; 276(35): 32591 - 32596. [Abstract] [Full Text] [PDF]