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Title:Proton relaxation studies of the interaction of small molecules with cytochrome P450
Author(s):Philson, Stephen Baker
Doctoral Committee Chair(s):Debrunner, Peter G.
Department / Program:Physics
Subject(s):proton relaxation
small molecule interactions
cytochrome P450
small molecule-proton complex
Abstract:The effect of ferric cytochrome P450 on the proton relaxation cam times of a variety of small molecules has been studied. In some cases, Fe-proton distances in the small molecule-protein complex are determined. For water protons the following results are obtained: The substrate (camphor) complex of P450, which is in a mainly high-spin ferric state, has little effect on H20 relaxation times, suggesting that the heme iron may be five-coordinate. On the other hand, the native, predominantly low-spin P450 has a large effect on the water. From the frequency and temperature dependence of the latter effect all the parameters associated with the relaxation are obtained. One or two exchangeable protons are clearly bound to an axial ligand of the iron. Among potential ligands, carboxylic acids, methionine, histidine, and cysteine can be definitely ruled out as causing the effect; this leaves hydroxyl, amine, and amide groups as possiblities, in addition to H0. 2Relaxation of the methyl protons of 2-methylpyridine by P450 indicates that the low-spin iron in the methylpyridine complex has an electron spin relaxation time TS of ~ 2 x 10-12 sec. This may be compared to the Ts for the native P450 of 5 x 10-10 sec, as derived from the frequency dependence of the water proton relaxation times. These TS values suggest that the relaxation effect of native P450 may be due not to the low-spin iron, but to a small high-spin fraction. Relaxation of the methyl protons of 5-exo-hydroxycamphor, the product of the reaction catalyzed by P450, is also studied. The effect of the protein is substantially smaller on the C-9 protons than on the others, suggesting orientation of the hydroxyl end of the molecule toward the heme iron and with an Fe-O distance of as little as 3 A. If camphor binds in the same way, it is ideally positioned for hydroxylation at the C-5 position by an Fe-02 complex. Relaxation studies of camphor protons are reported; however, their interpretation is difficult because of the complexity of the binding reaction. This complexity is evident from optical absorption measurements, which are also discussed.
Issue Date:1977
Genre:Dissertation / Thesis
Rights Information:1977 Stephen Baker Philson
Date Available in IDEALS:2011-07-01
Identifier in Online Catalog:252577

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