Previously, we isolated a selenate-and arsenate-reducing bacterium, designated strain SF-1, from seleniumcontaminated sediment and identified it as a novel species, Bacillus selenatarsenatis. B. selenatarsenatis strain SF-1 independently reduces selenate to selenite, arsenate to arsenite, and nitrate to nitrite by anaerobic respiration. To identify the genes involved in selenate reduction, 17 selenate reduction-defective mutant strains were isolated from a mutant library generated by random insertion of transposon Tn916. Tn916 was inserted into the same genome position in eight mutants, and the representative strain SF-1AM4 did not reduce selenate but did reduce nitrate and arsenate to the same extent as the wild-type strain. The disrupted gene was located in an operon composed of three genes designated srdBCA, which were predicted to encode a putative oxidoreductase complex by the BLASTX program. The plasmid vector pGEMsrdBCA, containing the srdBCA operon with its own promoter, conferred the phenotype of selenate reduction in Escherichia coli DH5␣, although E. coli strains containing plasmids lacking any one or two of the open reading frames from srdBCA did not exhibit the selenate-reducing phenotype. Domain structure analysis of the deduced amino acid sequence revealed that SrdBCA had typical features of membrane-bound and molybdopterin-containing oxidoreductases. It was therefore proposed that the srdBCA operon encoded a respiratory selenate reductase complex. This is the first report of genes encoding selenate reductase in Gram-positive bacteria.Selenium is the 34th element on the periodic table and has chemical properties resembling those of sulfur. It is obtained in limited ways, such as a by-product of the electric smelting of copper, and is an important material used in photoelectric devices, photosensitive drums used in dry copying, semiconductors, and the colorization and decolorization of glasses. Biologically, selenium is an essential element used for the synthesis of selenocysteine contained in selenoproteins, such as mammalian glutathione peroxidase and bacterial formate dehydrogenase. However, exposure to higher concentrations of selenium is toxic, and therefore it is important to understand how environmental selenium is controlled. Selenium has several oxidation states in the environment, i.e., selenate (ϩVI), selenite (ϩIV), elemental selenium (0), selenide (ϪII), and organic selenium (ϪII), and it is known that prokaryotes play a major role in its oxidation and reduction (28).The molecular mechanisms of selenate reduction have been analyzed in some bacteria. The selenate reductase complex of Thauera selenatis has been intensively studied biochemically and genetically. The selenate reductase complex of T. selenatis is a soluble periplasmic protein (20) that consists of three subunits, i.e., a catalytic subunit containing a molybdenum cofactor [Mo(V)], a subunit containing iron-sulfur clusters (one [3Fe-4S] cluster and three [4Fe-4S] clusters), and a subunit containing heme b (6, 25). The genes e...

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