CYP2E1 active site residues in substrate recognition sequence 5 identified by photoaffinity labeling and homology modeling.

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Collom SL, Jamakhandi AP, Tackett AJ, Radominska-Pandya A, Miller GP

CYP2E1 active site residues in substrate recognition sequence 5 identified by photoaffinity labeling and homology modeling.

Arch Biochem Biophys. 2007 Mar 1;459(1):59-69. doi: 10.1016/j.abb.2006.10.028. Epub 2006 Nov 2.

PubMed ID

17222385 [ View in PubMed

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Abstract

Despite its biological importance, our knowledge of active site structure and relevance of critical amino acids in CYP2E1 catalytic processes remain limited. In this study, we identified CYP2E1 active site residues using photoaffinity labeling with 7-azido-4-methylcoumarin (AzMC) coupled with a CYP2E1 homology model. In the absence of light, AzMC was an effective competitor against substrate p-nitrophenol oxidation by CYP2E1. Photoactivation of AzMC led to a concentration-dependent loss in CYP2E1 activity and structural integrity resulting from the modification of both heme and protein. The photo-labeling reaction degraded heme and produced a possible heme adduct. Probe incorporation into the protein occurred at multiple sites within substrate recognition sequence 5 (SRS-5). Based on a CYP2E1 homology model, we hypothesize AzMC labels SRS-5 residues, Leu363, Val364, and Leu368, in the active site. In addition, we propose a series of phenylalanines, especially Phe106, mediate contacts with the coumarin.

DrugBank Data that Cites this Article

Drug Enzymes

Drug	Enzyme	Kind	Organism	Pharmacological Action	Actions
Caffeine	Cytochrome P450 2E1	Protein	Humans	No	Substrate	Details