Biochemical Journal 2014-12-01

Cytochrome P450 system proteins reside in different regions of the endoplasmic reticulum.

Ji Won Park, James R Reed, Lauren M Brignac-Huber, Wayne L Backes

Index: Biochem. J. 464(2) , 241-9, (2014)

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Abstract

Cytochrome P450 (P450) function is dependent on the ability of these enzymes to successfully interact with their redox partners, NADPH-cytochrome P450 reductase (CPR) and cytochrome b5, in the endoplasmic reticulum (ER). Because the ER is heterogeneous in lipid composition, membrane microdomains with different characteristics are formed. Ordered microdomains are more tightly packed, and enriched in saturated fatty acids, sphingomyelin and cholesterol, whereas disordered regions contain higher levels of unsaturated fatty acids. The goal of the present study was to determine whether the P450 system proteins localize to different regions of the ER. The localization of CYP1A2, CYP2B4 and CYP2E1 within the ER was determined by partial membrane solubilization with Brij 98, centrifugation on a discontinuous sucrose gradient and immune blotting of the gradient fractions to identify ordered and disordered microdomains. CYP1A2 resided almost entirely in the ordered regions of the ER with CPR also localized predominantly to this region. CYP2B4 was equally distributed between the ordered and disordered domains. In contrast, CYP2E1 localized to the disordered membrane regions. Removal of cholesterol (an important constituent of ordered domains) led to the relocation of CYP1A2, CYP2B4 and CPR to the disordered regions. Interestingly, CYP1A1 and CYP1A2 localized to different membrane microdomains, despite their high degree of sequence similarity. These data demonstrate that P450 system enzymes are organized in specific membrane regions, and their localization can be affected by depletion of membrane cholesterol. The differential localization of different P450 in specific membrane regions may prov Related Materials Product # Image Description Molecular Formula Add to Cart SAB2501419 Anti-CYP2E1 antibody produced in goat, affinity isolated antibody, buffered aqueous solution pricing SAB1405688 Anti-CYP2E1 antibody produced in mouse, purified immunoglobulin, buffered aqueous solution pricing HPA009128 Anti-CYP2E1 antibody produced in rabbit, Prestige Antibodies® Powered by Atlas Antibodies, affinity isolated antibody, buffered aqueous glycerol solution pricing AV41424 Anti-CYP2E1 antibody produced in rabbit, affinity isolated antibody pricing HPA029564 Anti-CYP2E1 antibody produced in rabbit, Prestige Antibodies® Powered by Atlas Antibodies, affinity isolated antibody, buffered aqueous glycerol solution pricing 388866 Brij® 93, average Mn ~357 C22H44O3 pricing P6136 Brij® O10 C38H76O11 pricing 431281 Brij® O10, average Mn ~709 C38H76O11 pricing 436240 BRIJ® O20, average Mn ~1,150 C58H116O21 pricing C2155000 Cholesterol, European Pharmacopoeia (EP) Reference Standard C27H46O pricing PHR1533 Cholesterol, pharmaceutical secondary standard; traceable to USP, PhEur C27H46O pricing 47127-U Cholesterol solution, certified reference material, 10 mg/mL in chloroform C27H46O pricing 03610 Ethylenediaminetetraacetic acid, for complexometry, EDTA, ≥99.0% C10H16N2O8 pricing 03609 Ethylenediaminetetraacetic acid, BioUltra, ≥99.0% (KT) C10H16N2O8 pricing 39692 Ethylenediaminetetraacetic acid, TraceSELECT®, ≥99.995% C10H16N2O8 pricing 03620 Ethylenediaminetetraacetic acid, ≥98.0% (KT) C10H16N2O8 pricing 03690 Ethylenediaminetetraacetic acid solution, BioUltra, for molecular biology, pH 8.0, ~0.5 M in H2O C10H16N2O8 pricing 421332 MERPOL® OJ surfactant pricing SAB4100017 Monoclonal Anti-CYP2E1 antibody produced in mouse, culture supernatant pricing SAB4100018 Monoclonal Anti-CYP2E1 antibody produced in mouse, culture supernatant pricing P1636 Phenobarbital C12H12N2O3 pricing P-008 Phenobarbital solution, 1 mg/mL in methanol, ampule of 1 mL, certified reference material C12H12N2O3 pricing P5641 Polyoxyethylene (20) oleyl ether pricing 82620 Pyrazole, purum, ≥98.0% (GC) C3H4N2 pricing P56607 Pyrazole, 98% C3H4N2 pricing