title: A NapC/NirT-type cytochrome c (NrfH) is the mediator between the quinone pool and the cytochrome c nitrite reductase of Wolinella succinogenes.
abstract: Wolinella succinogenes can grow by anaerobic respiration with nitrate or nitrite using formate as electron donor. Two forms of nitrite reductase were isolated from the membrane fraction of W. succinogenes. One form consisted of a 58 kDa polypeptide (NrfA) that was identical to the periplasmic nitrite reductase. The other form consisted of NrfA and a 22 kDa polypeptide (NrfH). Both forms catalysed nitrite reduction by reduced benzyl viologen, but only the dimeric form catalysed nitrite reduction by dimethylnaphthoquinol. Liposomes containing heterodimeric nitrite reductase, formate dehydrogenase and menaquinone catalysed the electron transport from formate to nitrite; this was coupled to the generation of an electrochemical proton potential (positive outside) across the liposomal membrane. It is concluded that the electron transfer from menaquinol to the catalytic subunit (NrfA) of W. succinogenes nitrite reductase is mediated by NrfH. The structural genes nrfA and nrfH were identified in an apparent operon (nrfHAIJ) with two additional genes. The gene nrfA encodes the precursor of NrfA carrying an N-terminal signal peptide (22 residues). NrfA (485 residues) is predicted to be a hydrophilic protein that is similar to the NrfA proteins of Sulfurospirillum deleyianum and of Escherichia coli. NrfH (177 residues) is predicted to be a membrane-bound tetrahaem cytochrome c belonging to the NapC/NirT family. The products of nrfI and nrfJ resemble proteins involved in cytochrome c biogenesis. The C-terminal third of NrfI (902 amino acid residues) is similar to CcsA proteins from Gram-positive bacteria, cyanobacteria and chloroplasts. The residual N-terminal part of NrfI resembles Ccs1 proteins. The deduced NrfJ protein resembles the thioredoxin-like proteins (ResA) of Helicobacter pylori and of Bacillus subtilis, but lacks the common motif CxxC of ResA. The properties of three deletion mutants of W. succinogenes (DeltanrfJ, DeltanrfIJ and DeltanrfAIJ) were studied. Mutants DeltanrfAIJ and DeltanrfIJ did not grow with nitrite as terminal electron acceptor or with nitrate in the absence of NH4+ and lacked nitrite reductase activity, whereas mutant DeltanrfJ showed wild-type properties. The NrfA protein formed by mutant DeltanrfIJ seemed to lack part of the haem C, suggesting that NrfI is involved in NrfA maturation.
authors.affiliation: Institut für Mikrobiologie, Johann Wolfgang Goethe-Universität, Marie-Curie-Str. 9, D-60439 Frankfurt am Main, Germany.
authors.forename: J
authors.initials: J
authors.lastname: Simon
authors.forename: R
authors.initials: R
authors.lastname: Gross
authors.forename: O
authors.initials: O
authors.lastname: Einsle
authors.forename: P M
authors.initials: PM
authors.lastname: Kroneck
authors.forename: A
authors.initials: A
authors.lastname: Kröger
authors.forename: O
authors.initials: O
authors.lastname: Klimmek
mesh_terms: D000595:Amino Acid Sequence; D017124:Conserved Sequence; D003574:Cytochrome c Group / Q000235:genetics / Q000378:metabolism; D045263:Cytochromes a1; D003575:Cytochromes c1; D017353:Gene Deletion; D008081:Liposomes / Q000378:metabolism; D008969:Molecular Sequence Data; D005810:Multigene Family; D009154:Mutation; D009565:Nitrate Reductases / Q000235:genetics* / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism; D011809:Quinones / Q000378:metabolism*; D016968:Wolinella / Q000235:genetics / Q000378:metabolism*
chemical_list: D003574:Cytochrome c Group; D008081:Liposomes; D009573:Nitrites; D011809:Quinones; D045263:Cytochromes a1; D003575:Cytochromes c1; C017165:cytochrome c(3); D009565:Nitrate Reductases; C054690:nitrate reductase (cytochrome)