Cytochrome P-450 55A1 (P-450dNIR) acts as nitric oxide reductase employing NADH as the direct electron donor.
Cytochrome P-450dNIR (P-450dNIR), involved in the fungal denitrification by Fusarium oxysporum, was purified to homogeneity. The cytochrome P-450 (P-450) exhibited a potent nitric oxide (NO) reductase activity to form nitrous oxide (N2O) employing NADH but not NADPH as the sole effective electron donor. The apparent maximum turnover rate against NO was estimated as high as 31,500 min-1. The stoichiometry of the reaction between NO:NADH:N2O was 2:1:1. The reaction required neither an artificial electron-carrying mediator nor other proteinaceous components. An anaerobic incubation of ferric P-450dNIR.NO complex, but not of free ferric P-450dNIR, resulted in rapid reduction of the P-450, indicating that P-450dNIR.NO complex was reduced directly by NADH. Spectral changes during catalytic turnover indicated that decomposition of the ferrous P-450.NO complex might be rate-limiting. The reaction was not inhibited by carbon monoxide at all, suggesting that the free ferrous P-450 is not formed during turnover. On the basis of these results a possible reaction mechanism was considered. The present results demonstrated not only the unique reaction catalyzed by P-450, but also the first P-450-dependent reaction where the electron transport from NAD(P)H to P-450 is not supported by other components. Now that the physiological function of P-450 55A1 (P-450dNIR) has been found, we propose to alter the trivial name to P-450nor.
Nakahara
K
K
Institute of Applied Biochemistry, University of Tsukuba, Ibaraki, Japan.
Tanimoto
T
T
Hatano
K
K
Usuda
K
K
Shoun
H
H
D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003577:Cytochrome P-450 Enzyme System / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005670:Fusarium / Q000201:enzymology*; D007700:Kinetics; D008970:Molecular Weight; D009243:NAD / Q000378:metabolism*; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism*; D013053:Spectrophotometry; D013379:Substrate Specificity
D009243:NAD; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; C080610:nitric-oxide reductase (P450)