Purification and properties of trimethylamine N-oxide reductase from aerobic photosynthetic bacterium Roseobacter denitrificans.
Trimethylamine N-oxide (TMAO) reductase was purified from an aerobic photosynthetic bacterium Roseobacter denitrificans. The enzyme was purified from cell-free extract by ammonium sulfate fractionation, DEAE ion exchange chromatography, hydrophobic chromatography, and gel filtration. The purified enzyme was composed of two identical subunits with molecular weight of 90,000, as identified by SDS-polyacrylamide gel electrophoresis, containing heme c and a molybdenum cofactor. The molecular weight of the native enzyme determined by gel filtration was 172,000. The midpoint redox potential of heme c was +200 mV at pH 7.5. Absorption maxima appeared at 418,524, and 554 nm in the reduced state and 410 nm in the oxidized state. The enzyme reduced TMAO, nicotine acid N-oxide, picoline N-oxide, hydroxylamine, and bromate, but not dimethyl sulfoxide, methionine sulfoxide, chlorate, nitrate, or thiosulfate. Cytochrome c2 served as a direct electron donor. It probably catalyzes the electron transfer from cytochrome b-c1 complex to TMAO reductase. Cytochrome c552, another soluble low-molecular-weight cytochrome of this bacterium, also donated electrons directly to TMAO reductase.
Arata
H
H
Department of Biology, Faculty of Science, Kyushu University, Fukuoka.
Shimizu
M
M
Takamiya
K
K
D001419:Bacteria / Q000201:enzymology; D001420:Bacteria, Aerobic / Q000201:enzymology*; D001426:Bacterial Proteins; D003574:Cytochrome c Group / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006418:Heme / Q000031:analogs & derivatives / Q000032:analysis; D008970:Molecular Weight; D008982:Molybdenum / Q000032:analysis; D009247:NADH, NADPH Oxidoreductases / Q000302:isolation & purification* / Q000378:metabolism; D010084:Oxidation-Reduction; D000587:Oxidoreductases Acting on CH-NH Group Donors; D010788:Photosynthesis; D012680:Sensitivity and Specificity; D012967:Sodium Dodecyl Sulfate; D013053:Spectrophotometry
D001426:Bacterial Proteins; D003574:Cytochrome c Group; C083951:cytochrome c553; C031690:heme C; D012967:Sodium Dodecyl Sulfate; D006418:Heme; D008982:Molybdenum; C017167:cytochrome C(551); C022302:cytochrome C-552; C021567:methylamine dehydrogenase; D000587:Oxidoreductases Acting on CH-NH Group Donors; D009247:NADH, NADPH Oxidoreductases