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Multiple substrate binding by cytochrome P450 3A4: estimation of the number of bound substrate molecules.

Kapelyukh, Yury, Paine, Mark ORCID:, Maréchal, Jean-Didier, Sutcliffe, Michael J., Wolf, C. Roland and Roberts, Gordon C. K. (2008) 'Multiple substrate binding by cytochrome P450 3A4: estimation of the number of bound substrate molecules.'. Drug metabolism and disposition: the biological fate of chemicals, Vol 36, Issue 10, pp. 2136-44.

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Cytochrome P450 3A4, a major drug-metabolizing enzyme in man, is well known to show non-Michaelis-Menten steady-state kinetics for a number of substrates, indicating that more than one substrate can bind to the enzyme simultaneously, but it has proved difficult to obtain reliable estimates of exactly how many substrate molecules can bind. We have used a simple method involving studies of the effect of large inhibitors on the Hill coefficient to provide improved estimates of substrate stoichiometry from simple steady-state kinetics. Using a panel of eight inhibitors, we show that at least four molecules of the widely used CYP3A4 substrate 7-benzyloxyquinoline can bind simultaneously to the enzyme. Computational docking studies show that this is consistent with the recently reported crystal structures of the enzyme. In the case of midazolam, which shows simple Michaelis-Menten kinetics, the inhibitor effects demonstrate that two molecules must bind simultaneously, consistent with earlier evidence, whereas for diltiazem, the experiments provide no evidence for the binding of more than one molecule. The consequences of this "inhibitor-induced cooperativity" for the prediction of pharmacokinetics and drug-drug interactions are discussed.

Item Type: Article
Subjects: QV Pharmacology > QV 38 Drug action.
Faculty: Department: Groups (2002 - 2012) > Vector Group
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Depositing User: Users 183 not found.
Date Deposited: 25 Aug 2010 10:53
Last Modified: 06 Feb 2018 13:01


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