Intermediates in the reaction of substrate‐free cytochrome P450cam with peroxy acetic acid

FEBS Letters

Trautwein, A.X.³  Mandon, D.²  Jung, C.¹  Weiss, R.²  Schünemann, V.³ 

[1] Max-Delbrück-Center for Molecular Medicine, 13092 Berlin, Germany;Laboratoire de Cristallochimie, UMR 7513, Université Louis Pasteur, 67070 Strasbourg, France;Institute of Physics, Medical University, 23538 Lübeck, Germany

关键词: Cytochrome P450cam;    Peroxy acetic acid;    Mössbauer spectrum;    P450cam;    cytochrome P450cam (CYP101) from Pseudomonas putida which catalyzes the hydroxylation of 1R-camphor;    DTT;    DL-dithiothreitol;    DEAE;    diethylaminoethyl;   

DOI  :  10.1016/S0014-5793(00)01886-X

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Freeze-quenched intermediates of substrate-free cytochrome ⁵⁷Fe-P450_cam in reaction with peroxy acetic acid as oxidizing agent have been characterized by EPR and Mössbauer spectroscopy. After 8 ms of reaction time the reaction mixture consists of ∼90% of ferric low-spin iron with g-factors and hyperfine parameters of the starting material; the remaining ∼10% are identified as a free radical (S′=1/2) by its EPR and as an iron(IV) (S=1) species by its Mössbauer signature. After 5 min of reaction time the intermediates have disappeared and the Mössbauer and EPR-spectra exhibit 100% of the starting material. We note that the spin-Hamiltonian analysis of the spectra of the 8 ms reactant clearly reveals that the two paramagnetic species, e.g. the ferryl (iron(IV)) species and the radical, are not exchanged coupled. This led to the conclusion that under the conditions used, peroxy acetic acid oxidized a tyrosine residue (probably Tyr-96) into a tyrosine radical (Tyr-96), and the iron(III) center of substrate-free P450_cam to iron(IV).

【 授权许可】

Unknown