High-throughput functional screening of steroid substrates with wild-type and chimeric P450 enzymes.

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Urban P, Truan G, Pompon D

High-throughput functional screening of steroid substrates with wild-type and chimeric P450 enzymes.

Biomed Res Int. 2014;2014:764102. doi: 10.1155/2014/764102. Epub 2014 Aug 26.

PubMed ID

25243177 [ View in PubMed

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Abstract

The promiscuity of a collection of enzymes consisting of 31 wild-type and synthetic variants of CYP1A enzymes was evaluated using a series of 14 steroids and 2 steroid-like chemicals, namely, nootkatone, a terpenoid, and mifepristone, a drug. For each enzyme-substrate couple, the initial steady-state velocity of metabolite formation was determined at a substrate saturating concentration. For that, a high-throughput approach was designed involving automatized incubations in 96-well microplate with sixteen 6-point kinetics per microplate and data acquisition using LC/MS system accepting 96-well microplate for injections. The resulting dataset was used for multivariate statistics aimed at sorting out the correlations existing between tested enzyme variants and ability to metabolize steroid substrates. Functional classifications of both CYP1A enzyme variants and steroid substrate structures were obtained allowing the delineation of global structural features for both substrate recognition and regioselectivity of oxidation.

DrugBank Data that Cites this Article

Drug Enzymes

Drug	Enzyme	Kind	Organism	Pharmacological Action	Actions
Testosterone	Cytochrome P450 1A1	Protein	Humans	Unknown	Substrate	Details