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Epoxidation of benzo[a]pyrene-7,8-dihydrodiol by human CYP1A1 in reconstituted membranes

Item Type:Article
Title:Epoxidation of benzo[a]pyrene-7,8-dihydrodiol by human CYP1A1 in reconstituted membranes
Creators Name:Kisselev, P. and Schwarz, D. and Platt, K.L. and Schunck, W.H. and Roots, I.
Abstract:Human cytochrome P4501A1 (CYP1A1) is one of the key enzymes in the bioactivation of environmental pollutants such as benzo[a]pyrene (B[a]P) and other polycyclic aromatic hydrocarbons. To evaluate the effect of membrane properties and distinct phospholipids on the activity of human CYP1A1 purified insect cell-expressed human CYP1A1 and of human NADPH-P450 reductase were reconstituted into phospholipid vesicle membranes. Conversion rates of up to 36 pmol·min−1·pmol−1 CYP1A1 of the enantiomeric promutagens (–)- and (+)-trans-7,8-dihydroxy-7,8-dihydro-B[a]P (7,8-diol) to the genotoxic diolepoxides were achieved. The highest rates were obtained when negatively charged lipids such as phosphatidylserine and phosphatidylinositol and/or nonbilayer phospholipids such as phosphatidylethanolamine were present in the membrane together with neutral lipids. Both Vmax and Km values were changed. This suggests a rather complex mechanism of stimulation which might include altered substrate binding as well as more effective interaction between CYP1A1 and NADPH-P450 reductase. Furthermore, the ratio of r-7,t-8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydro-B[a]P (DE2) to r-7,t-8-dihydroxy-c-9,10-epoxy-7,8,9,10-tetrahydro-B[a]P (DE1) formed from (–)-7,8-diol was significantly increased by the introduction of anionic lipids, but not by that of nonbilayer lipids. Thus, charged lipids affect the stereoselectivity of the epoxidation by leading to the formation of a larger amount of the ultimate mutagen DE2 than of DE1, which is far less carcinogenic. These data suggest that membrane properties such as negative charge and nonbilayer phase propensity are important for the efficiency and selectivity of enzymatic function of human CYP1A1.
Keywords:Human Cytochrome P4501A1, Vesicle Reconstitution, Epoxidation, Benzo[a]pyrene, Benzo[a]pyrene-7, 8diol
Source:European Journal of Biochemistry
ISSN:0014-2956
Publisher:Blackwell Publishing (U.K.)
Volume:269
Number:7
Page Range:1799-1805
Date:April 2002
Official Publication:https://doi.org/10.1046/j.1432-1033.2002.02848.x
PubMed:View item in PubMed

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