Summary: strains capable of dissimilatory nitrous oxide reduction (Nos) carry a gene cluster on a 10.1 kb RI fragment of the megaplasmid near the genes. These genes are arranged in three complementation groups and the 10.1 kb RI fragment is sufficient to confer Nos activity to strains lacking such activity. An overlapping dIII fragment containing the genes but missing a 0.6 kb RI downstream segment was found incapable of imparting Nos activity to strains unable to reduce nitrous oxide, suggesting the presence of other gene(s) in this region. Tn5 introduced near the dIII site resulted in mutants with a Nos phenotype. Complete sequence analysis of showed that it was well-conserved with respect to that of . Two previously unreported genes downstream of in were also revealed. Contiguous with was a sequence showing 63% identity with the ORFL protein of . It appeared to be in the same operon as and was predicted to encode a membrane lipoprotein similar to the putative NosL of . Unlike the latter protein, however, amino acid sequences typical of metal-binding sites and cysteine residues indicative of the active site of protein disulphide isomerase were absent in the predicted NosL of . The Tn5 mutations resulting in a Nos phenotype were localized within a 966 nucleotide gene 31 nucleotides downstream of with the same orientation. The new gene, , was determined to be in a separate complementation group. It encoded a periplasmic protein with homology in the C-terminal domain with RnfF of and with a hypothetical Escherichia coli protein, YOJK. It was concluded that there are seven genes constituting the cluster in . They are organized in four complementation groups and in the same orientation, spanning a distance of about 9 kb on the megaplasmid.


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