Conformational changes near the cytochrome P450 active site upon binding of two different ligands.
It is shown that a stable nitroxyl radical, 4-cyano-2,2,6,6-tetramethylpiperidine-1-oxyl, forms a complex with cytochrome P4502B4 by analogy with the second type substrates by joining directly to pentacoordinate heme iron. The bound radical is inaccessible to water-soluble paramagnetic ions, which confirms its localization in a hydrophobic pocket near the heme. Benzphetamine and N,N-dimethylaniline, the first-type nonpolar substrates, induce conformational changes of the spin-labeled hemoprotein which are evidently accompanied by an increase in the volume of the pocket resulting in emergence of contact with aqueous phase, and the heme-bound spin label becomes accessible to water-soluble paramagnetics. In this case potassium ferricyanide broadens the spin-labeled cytochrome signal and, as a result, lowers the amplitudes of the spectral components. Similar changes were registered at non-micellar concentrations of nonionic detergent Emulgen 913, whose activating effect on hydroxylation reactions is associated, as we showed previously, with its presence in the CYP2B4 active site simultaneously with substrates.
Myasoedova
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KN
Semenov Institute of Chemical Physics, Russian Academy of Sciences, Moscow, Russia. KseniaMyasoedova@gmail.com
Timofeev
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KN
D000814:Aniline Compounds / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases / Q000737:chemistry* / Q000378:metabolism; D001589:Benzphetamine / Q000737:chemistry / Q000378:metabolism; D020134:Catalytic Domain; D000072467:Cytochrome P450 Family 2; D006418:Heme / Q000737:chemistry / Q000378:metabolism; D008024:Ligands; D008862:Microsomes, Liver / Q000737:chemistry / Q000201:enzymology*; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D013379:Substrate Specificity
D000814:Aniline Compounds; D008024:Ligands; D001589:Benzphetamine; D006418:Heme; C015157:N,N-dimethylaniline; D001189:Aryl Hydrocarbon Hydroxylases; D000072467:Cytochrome P450 Family 2; C465241:cytochrome P-450 CYP2B4 (rabbit)