Submitted to: Bioremediation and Sustainable Environmental Technologies
Publication Type: Abstract Only
Publication Acceptance Date: 8/31/2013
Publication Date: 8/31/2013
Citation: Hunter, W.J., Manter, D.K. 2013. Rhizobium selenitireducens proteins involved in the reduction of selenite to elemental selenium. Bioremediation and Sustainable Environmental Technologies. 1.
Technical Abstract: Microbial based bioremediation barriers can remove the metalloid selenite (SeO3–2) from flowing groundwater. The organisms associated with the process include microorganisms from within the bacterial and archaeal domains that can reduce soluble SeO3–2 to the insoluble and reddish-colored elemental selenium (Se0). However, the enzymatic processes associated with SeO3–2 reduction are poorly understood. In a few microorganisms SeO3–2 reduction serves a respiratory function but this is not the case in most microorganisms where reduction may serve as a means of reducing the toxicity of SeO3–2. In Rhizobium selenitireducens a nitrite reductase has been shown to reduce SeO3–2 but other proteins are also involved; this study investigated two additional proteins. Cultures of R. selenitireducens were grown in media containing Na2SeO3 to induce reductase activity. When proteins from these cells, partially purified via ammonium sulfate precipitation, were run on native electrophoresis gels and the gels incubated with SeO3–2 reddish-orange bands of precipitated Se0 formed indicating the presence of SeO3–2 reductase activity. Two active proteins were detected, one utilized NADH as an electron donor and had an Rf of 0.78 while the second utilized NADPH and had an Rf of 0.65. The NADPH band reduced tellurite as well as SeO3–2 while the NADH dependent band did not reduce tellurite. Both bands were cut from the gel and analyzed for peptides via LCMSMS. Peptides associated with an NADH:flavin oxidoreductase, with Sinorhizobium medicae NADH:flavin oxidoreductase (YP_001326930) providing the strongest match in the GenBank database, were detected. YP_001326930 is part of a large family of proteins that contain an “old yellow enzyme” (OYE) like flavin binding domain; a name derived from the first flavin-dependent enzyme identified. OYE enzymes may not have a single specific substrate rather, due in part to an active site that has a large and accessible binding pocket, enzymes belonging to this family are often active on a wide range of substrates and are often involved in protecting cells from oxidative stress. This is the first report of an OYE enzyme being involved in SeO3–2 reduction though a T. scotoductus chromate reductase also belongs to the OYE protein family. The LCMSMS results failed to identify a match with the NADPH band.