Preliminary studies showed that the periplasmic nitrate reductase (Nap) of Rhodobacter sphaeroides and the membrane-bound nitrate reductases of Escherichia coli are able to reduce selenate and tellurite in vitro with benzyl viologen as an electron donor. In the present study, we found that this is a general feature of denitrifiers. Both the periplasmic and membrane-bound nitrate reductases of Ralstonia eutropha, Paracoccus denitrificans, and Paracoccus pantotrophus can utilize potassium selenate and potassium tellurite as electron acceptors. In order to characterize these reactions, the periplasmic nitrate reductase of R. sphaeroides f. sp. denitrificans IL106 was histidine tagged and purified. The V max and K m were determined for nitrate, tellurite, and selenate. For nitrate, values of 39 mol ⅐ min ؊1 ⅐ mg ؊1 and 0.12 mM were obtained for V max and K m , respectively, whereas the V max values for tellurite and selenate were 40-and 140-fold lower, respectively. These low activities can explain the observation that depletion of the nitrate reductase in R. sphaeroides does not modify the MIC of tellurite for this organism.Selenium is part of the amino acid selenocysteine present in numerous enzymes and is essential to all living cells (42). Furthermore, selenium can help prevent cancer and other diseases (11). However, at high concentrations this compound, predominantly in the form of selenate and selenite oxyanions, is toxic and can cause some environmental problems; for example, in the San Joaquin Valley in central California, bird malformations due to selenium have been reported (28). Tellurium is not an essential element and is relatively rare in the environment, but it can be found at high concentrations near waste discharge sites. It is also extremely toxic, and the MIC for Escherichia coli is approximately 2 g of potassium tellurite per ml (3). Nevertheless, some gram-negative organisms are resistant to potassium tellurite (24). Different explanations for resistance have been proposed; these include exclusion, increased efflux, and reduction to the less toxic metallic form. Several genetic determinants have been shown to confer tellurite resistance (15,27,44,45,48,50). Although physiological functions can be attributed to some of these determinants (for example, tpm and tehB, which exhibit homology with methyl transferases [1,8,19] and arsRDABC, which encodes an oxyanion efflux transporter [45]), most of them exhibit no similarity to each other or to the locus encoding any enzyme whose function is known. Therefore, the mechanisms which allow these loci to confer resistance remain largely unknown (46). When reduction occurs, intracellular deposition of tellurium can be observed, and bacteria form black colonies (20, 24). Resistance to selenium oxides is also partially attributed to reduction and accumulation of the red amorphous Se 0 form in the cell (12, 14; M. Bébien, J.-P. Chauvin, J.-M. Adriano, S.Grosse, and A. Verméglio, submitted for publication). For instance, the photosynthetic bacterium Rhodobacte...
