کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1318166 | 1499482 | 2006 | 11 صفحه PDF | دانلود رایگان |
Previously, we reported spectroscopic properties of cytochrome P450cam compound I, (ferryl iron plus a porphyrin π-cation radical (FeIV = O/Por+)), as well as compound ES (FeIV = O/Tyr) in reactions of substrate-free ferric enzyme with m-chloroperbenzoic acid [T. Spolitak, J.H. Dawson, D.P. Ballou, J. Biol. Chem. 280 (2005) 20300-9]. Compound ES arises by intramolecular electron transfer from nearby tyrosines to the porphyrin π-cation radical of Compound I, and has been characterized by rapid-freeze-quench-Mössbauer/EPR spectroscopy; the tyrosyl radical was assigned to Tyr96 for wild type or to Tyr75 for the Tyr96Phe variant [V. Schünemann, F. Lendzian, C. Jung, J. Contzen, A.L. Barra, S.G. Sligar, A.X. Trautwein, J. Biol. Chem. 279 (2004) 10919–10930]. Here we report rapid-scanning stopped-flow studies of the reactions of peracids with substrate-free ferric Y75F, Y96F, and Y96F/Y75F P450cam variants, showing how these active site changes influence electron transfer from nearby tyrosines and affect formation of intermediates. Curiously, rates of generation of Compounds I and ES for both single mutants were not very different from wild type. Contrasting with the earlier EPR results, the Y96F/Y75F variant was also shown to form an ES-like species, but more slowly. When substrate is not present, or is improperly bound, compound I rapidly converts to compound ES, which can be reduced to form H2O and ferric P450, thus avoiding the modification of nearby protein groups or release of reactive oxygen species.
Journal: Journal of Inorganic Biochemistry - Volume 100, Issue 12, December 2006, Pages 2034–2044