PMID:7508388

Citation	Zumft, WG, Braun, C and Cuypers, H (1994) Nitric oxide reductase from Pseudomonas stutzeri. Primary structure and gene organization of a novel bacterial cytochrome bc complex. Eur. J. Biochem. 219:481-90
Abstract	Nitric oxide (NO) reductase is an integral membrane component of the anaerobic respiratory chain of Pseudomonas stutzeri that transforms nitrate to dinitrogen (denitrification). The enzyme catalyzes the reduction of NO to nitrous oxide. The structural genes for the NO reductase complex, norC and norB, were sequenced and their organization established by primer extension and Northern blot analysis. The norCB genes encoding the cytochrome c and cytochrome b subunits of the enzyme are contiguous and transcribed as a single 2.0-kb transcript. The promoter region has a canonical recognition motif for the transcriptional activator protein Fnr, centered at -40.5 nucleotides from the initiation site of transcription. No similarity of the derived gene products to known cytochromes of b- or c-type was found in a data bank search. Post-translational processing of the two subunits was limited to the removal of the terminal methionine to leave an N-terminal serine in either subunit. The mature cytochrome c subunit (16508Da, 145 residues) is predicted to be a bitopic protein with a single membrane anchor. The mature cytochrome b subunit (53006Da, 473 residues) is a putatively polytopic, strongly hydrophobic membrane-bound protein with 12 potential transmembrane segments. Several histidine and proline residues were identified with potentially structural and/or functional importance. Mutational inactivation of NO reductase by deletion of norB or the norCB genes affected strongly the in vivo activity of respiratory nitrite reductase (cytochrome cd1), but to a much lesser extent the expression level of this enzyme. In turn, mutational inactivation of the structural gene for cytochrome cd1, nirS, or loss of in vivo nitrite reduction by mutation of the nirT gene, encoding a presumed tetraheme cytochrome, lowered the expression level of NO reductase to 5-20%, but hardly its catalytic activity. The cellular concentration of NO reductase increased again on restoration of nitrite reduction in the nirS::Tn5 mutant MK202 by complementation with nirS or with the heterologous nirK gene, encoding the Cu-containing nitrite reductase from Pseudomonas aureofaciens. Thus, NO may be required as an inducer for its own reductase. Our results show that the nitrite-reducing system and the NO-reducing system are not operating independently from each other but are interlaced by activity modulation and regulation of enzyme synthesis.
Links	PubMed
Keywords	Amino Acid Sequence; Base Sequence; Blotting, Southern; DNA, Bacterial/isolation & purification; DNA, Bacterial/metabolism; Electron Transport Complex III/biosynthesis; Electron Transport Complex III/genetics; Electron Transport Complex III/isolation & purification; Gene Deletion; Gene Expression; Genes, Bacterial; Molecular Sequence Data; Molecular Weight; Oxidoreductases/biosynthesis; Oxidoreductases/genetics; Oxidoreductases/isolation & purification; Pseudomonas/enzymology; Pseudomonas/genetics; Pseudomonas/growth & development; RNA, Bacterial/isolation & purification; RNA, Bacterial/metabolism; Restriction Mapping; Transcription, Genetic
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
PSEST:NORB		GO:0016966: nitric oxide reductase activity	ECO:0000315:		F		Figure 8. Deletion-replacement mutagenesis of the gene norB resulted in the loss of NO reduction of P stutzeri strain MK321.	complete CACAO 2488
PSEST:NORB	enables	GO:0016966: nitric oxide reductase activity	ECO:0000315: mutant phenotype evidence used in manual assertion		F		Seeded From UniProt	complete

References

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