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A Mo6+-reducing bacterium (strain 48), which grew on medium supplemented with 200 mM Mo6+, was isolated from stream water obtained from Chengkau, Malaysia. The chemical properties of strain 48 conform to the characteristics of Enterobacter cloacae. Under anaerobic conditions in the glucose-yeast extract medium containing phosphate ion (2.9 mM) and Mo6+ (10 mM), the bacterium reduced Mo6+ to form molybdenum blue. Approximately 27% of Mo6+ added to the medium was reduced after 28 h of cultivation. The reduction of Mo6+ with glucose as an electron donor was strongly inhibited by iodoacetic acid, sodium fluoride, and sodium cyanide, suggesting an involvement of the glycolytic pathway and electron transport in Mo6+ reduction. NADH and N,N,N',N'-tetramethyl-p-phenylenediamine served as electron donors for Mo6+ reduction. When NADH was used as an electron donor, at first cytochrome b in the cell extract was reduced, and then molybdenum blue was formed. Sodium cyanide strongly inhibited Mo6+ reduction by NADH (5 mM) but not the reduction of cytochrome b in the cell extract, suggesting that the reduced component of the electron transport system after cytochrome b serves as an electron donor for Mo6+ reduction. Both ferric and stannous ions strongly enhanced the activity of Mo6+ reduction by NADH.

A sulfur:ferric ion oxidoreductase that utilizes ferric ion (Fe3+) as an electron acceptor of elemental sulfur was purified from iron-grown Thiobacillus ferrooxidans to an electrophoretically homogeneous state. Under anaerobic conditions in the presence of Fe3+, the enzyme reduced 4 mol of Fe3+ with 1 mol of elemental sulfur to give 4 mol of Fe2+ and 1 mol of sulfite, indicating that it corresponds to a ferric ion-reducing system (T. Sugio, C. Domatsu, O. Munakata, T. Tano, and K. Imai, Appl. Environ. Microbiol. 49:1401-1406, 1985). Under aerobic conditions, sulfite, but not Fe2+, was produced during the oxidation of elemental sulfur by this enzyme because the Fe2+ produced was rapidly reoxidized chemically by molecular oxygen. The possibility that Fe3+ serves as an electron acceptor under aerobic conditions was ascertained by adding o-phenanthroline, which chelates Fe2+, to the reaction mixture. Sulfur:ferric ion oxidoreductase had an apparent molecular weight of 46,000, and it is composed of two identical subunits (Mr = 23,000) as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Sulfur oxidation by this enzyme was absolutely dependent on the presence of reduced glutathione. The enzyme had an isoelectric point and a pH optimum at pH 4.6 and 6.5, respectively. Almost all the activity of sulfur:ferric ion oxidoreductase was observed in the osmotic shock fluid of the cells, suggesting that it was localized in the periplasmic space of the cells.

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Sulfite:ferric ion oxidoreductase in the plasma membrane of Thiobaciusferrooxidans AP19-3 was purified to an electrophoretically homogeneous state. The enzyme had an apparent molecular weight of 650,000 and was composed of two subunits (M1s, 61,000 and 59,000) as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The Michaelis constants of sulfite:ferric ion oxidoreductase for Fe3' and sulfite ions were 1.0 and 0.071 mM, respectively. Sulfite:ferric ion oxidoreductase suffered from end product inhibition by 1 mM Fe2+.