Cytochrome P-460 of Nitrosomonas europaea: further purification and further characterization.
Cytochrome P-460 of Nitrosomonas europaea [Erickson, R.H. and Hooper, A.B. (1972) Biochim. Biophys. Acta 275, 231-244] was further purified to an electrophoretically homogeneous state. The cytochrome molecule was composed of three molecules of subunits with Mr of 17,300-18,500, and contained three atoms of iron, which seemed to be heme iron, and six cysteine residues, but did not contain nonheme iron or inorganic sulfide. The cytochrome showed absorption peaks at 460 and 688 nm with a broad shoulder at 635 nm in the reduced form. The ESR spectrum of ferricytochrome P-460 showed signals at g = 5.91, 5.63, and 1.99, indicating that the protein was a high spin hemoprotein. The heme of the cytochrome was not cleaved by the methods which were available for cleavage of heme c. The pyridine ferrohemochrome of the hemoprotein did not show the distinct alpha and beta peaks which are shown by the ferrohemochromes of many other cytochromes so far known. The N-terminal amino acid sequence of cytochrome P-460 differed from that of hydroxylamine oxidoreductase. Therefore, cytochrome P-460 did not seem to be the solubilized P-460 moiety of hydroxylamine oxidoreductase, in agreement with the finding by D.J. Miller et al. [J. Gen. Microbiol. 130, 3049-3054 (1984)]. However, cytochrome P-460 had several enzymatic activities which hydroxylamine oxidoreductase showed. Although most of the activities of the cytochrome were lower than the corresponding activities of the oxidoreductase, the hydroxylamine-cytochrome c-552 reductase activity of the cytochrome was about 5-times as high as that of the oxidoreductase.
Numata
M
M
Department of Life Science, Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology.
Saito
T
T
Yamazaki
T
T
Fukumori
Y
Y
Yamanaka
T
T
D000595:Amino Acid Sequence; D003580:Cytochromes / Q000737:chemistry / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D006418:Heme / Q000737:chemistry; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009606:Nitrosomonas / Q000187:drug effects / Q000201:enzymology*; D010088:Oxidoreductases / Q000378:metabolism; D012394:Rosaniline Dyes; D012977:Sodium Nitrite / Q000494:pharmacology
D003580:Cytochromes; D012394:Rosaniline Dyes; D006418:Heme; D010088:Oxidoreductases; C021401:hydroxylamine dehydrogenase; D012977:Sodium Nitrite; C004692:coomassie Brilliant Blue