title: Electron transfer to nitrite reductase of Rhodobacter sphaeroides 2.4.3: examination of cytochromes c2 and cY.
abstract: The role of cytochrome c(2), encoded by cycA, and cytochrome c(Y), encoded by cycY, in electron transfer to the nitrite reductase of Rhodobacter sphaeroides 2.4.3 was investigated using both in vivo and in vitro approaches. Both cycA and cycY were isolated, sequenced and insertionally inactivated in strain 2.4.3. Deletion of either gene alone had no apparent effect on the ability of R. sphaeroides to reduce nitrite. In a cycA-cycY double mutant, nitrite reduction was largely inhibited. However, the expression of the nitrite reductase gene nirK from a heterologous promoter substantially restored nitrite reductase activity in the double mutant. Using purified protein, a turnover number of 5 s(-1) was observed for the oxidation of cytochrome c(2) by nitrite reductase. In contrast, oxidation of c(Y) only resulted in a turnover of approximately 0.1 s(-1). The turnover experiments indicate that c(2) is a major electron donor to nitrite reductase but c(Y) is probably not. Taken together, these results suggest that there is likely an unidentified electron donor, in addition to c(2), that transfers electrons to nitrite reductase, and that the decreased nitrite reductase activity observed in the cycA-cycY double mutant probably results from a change in nirK expression.
authors.affiliation: Department of Microbiology, Wing Hall, Cornell University, Ithaca, NY 14850-8101, USA.
authors.forename: William P
authors.initials: WP
authors.lastname: Laratta
authors.affiliation: Department of Microbiology, Wing Hall, Cornell University, Ithaca, NY 14850-8101, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Nanaszko
authors.affiliation: Department of Microbiology, Wing Hall, Cornell University, Ithaca, NY 14850-8101, USA.
authors.forename: James P
authors.initials: JP
authors.lastname: Shapleigh
mesh_terms: D000595:Amino Acid Sequence; D003574:Cytochrome c Group / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D045362:Cytochromes c2 / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004579:Electron Transport; D017353:Gene Deletion; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D050901:Nitrate Reductase / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism; D010084:Oxidation-Reduction; D012242:Rhodobacter sphaeroides / Q000235:genetics / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
chemical_list: D003574:Cytochrome c Group; D004269:DNA, Bacterial; D009573:Nitrites; C080693:cytochrome Cy; D045362:Cytochromes c2; D050901:Nitrate Reductase