Electron spin echo envelope modulation studies of the semiquinone anion radical of cholesterol oxidase from Brevibacterium sterolicum

FEBS Letters

Medina, Milagros²  Vrielink, Alice¹  Cammack, Richard² 

[1] Department of Biochemistry, McGill University, 3655 Drummond Street, Montreal, Que. H3G 1Y6, Canada;Centre for the Study of Metals in Biology and Medicine, King's College, Campden Hill Road, London W8 7AH, UK

关键词: Cholesterol oxidase;    Brevibacterium sterolicum;    Electron spin echo envelope modulation;    Flavoprotein semiquinone;    Hyperfine interaction;    Pulsed EPR spectroscopy;    EPR;    electron paramagnetic resonance;    ENDOR;    electron-nuclear double resonance;    ESEEM;    electron spin echo envelope modulation;    GMC;    glucose-methanol-choline;    NQR;    nuclear quadrupole resonance;    η;    asymmetry parameter;    e2qQ;    quadrupole coupling constant;    A;    hyperfine coupling constant;    ν i;    nuclear Zeeman frequency;   

DOI  :  10.1016/S0014-5793(96)01372-5

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The electron spin echo envelope modulation (ESEEM) technique of pulsed EPR spectroscopy was applied to the anionic semiquinone of the cholesterol oxidase flavin cofactor, formed when the enzyme was photoreduced in the presence of 5-deazariboflavin and EDTA. Fourier transforms of the three-pulse ESEEM spectra showed the presence of ¹⁴N nuclei magnetically coupled to the paramagnet. In ²H₂O buffer the surroundings of the flavin ring were shown to be accessible to solvent exchange, with a deuterium population in close proximity to the paramagnetic centre. Upon binding of the pseudosubstrate, dehydroisoandrosterone, subtle changes were observed in the coupling to nitrogen nuclei, which are interpreted as changes in the electron density distribution of the flavin ring system. The results are discussed in terms of the three-dimensional structure reported for the protein and the flavin ring architecture.

【 授权许可】

Unknown