Molecular characterization of the nitrite-reducing system of Staphylococcus carnosus

Characterization of a nitrite reductase-negative Staphylococcus carnosus Tn 917 mutant led to the identification of the nir operon, which encodes NirBD , the dissimilatory NADH-dependent nitrite reductase; SirA, the putative ox idase and chelatase, and SirB, the uroporphyrinogen III methylase, both of which are necessary for biosynthesis of the siroheme prosthetic group; and NirR, which revealed no convincing similarity to proteins with known functi ons. We suggest that NirR is essential for nir promoter activity. In the ab sence of NirR, a weak promoter upstream of sirA seems to drive transcriptio n of sirA, nirB, nirD, and sirB in the stationary-growth phase. In primer e xtension experiments one predominant and several weaker transcription start sites were identified in the nir promoter region. Northern blot analyses i ndicated that anaerobiosis and nitrite are induction factors of the nil ope ron: cells grown aerobically,vith nitrite revealed small amounts of full le ngth transcript whereas cells grown anaerobically with or without nitrite s howed large amounts of full-length transcript. Although a transcript is det ectable, no nitrite reduction occurs in cells grown aerobically with nitrit e, indicating an additional oxygen-controlled step at the level of translat ion, enzyme folding, assembly, or insertion of prosthetic groups. The nitri te-reducing activity expressed during anaerobiosis is switched off reversib ly when the oxygen tension increases, most likely due to competition for el ectrons with the aerobic respiratory chain. Another gene, nirC, is located upstream of the nir operon, nirC encodes a putative integral membrane-spann ing protein of unknown function. A nirC mutant showed no distinct phenotype .