Rotation of cytochrome P-450. I. Investigations of protein-protein interactions of cytochrome P-450 in phospholipid vesicles and liver microsomes.

This study aimed on exploration of the system-wide effects of the alcohol-induced increase in the content of cytochrome P450 2E1 (CYP2E1) in the human liver on drug metabolism. Using membrane incorporation of purified CYP2E1 modified with photoreactive crosslinkers benzophenone-4-maleimide (BPM) and 4-(N-succinimidylcarboxy)benzophenone (BPS), we explored the array of its protein-protein interactions (proteome) in human liver microsomes (HLM) with chemical cross-linking mass spectrometry (CXMS). Exposure of bait-incorporated HLM samples to light was followed by isolation of the His-tagged bait protein and its cross-linked aggregates on Ni-NTA agarose. Analyzing the individual bands of SDS-PAGE slabs of thereby isolated protein with the toolset of untargeted proteomics, we detected the cross-linked dimeric and trimeric complexes of CYP2E1 with other drug-metabolizing enzymes. Among the most extensively cross-linked partners of CYP2E1 are cytochromes P450 2A6, 3A4, 2C9, and 4A11. We also detected the conjugates of CYP2E1 with UDP-glucuronosyltransferases (UGTs) 1A6, 1A9, 2B4, 2B15, and 2B17. These results demonstrate the exploratory power of the proposed CXMS strategy and corroborate the concept of tight functional integration in the human drug-metabolizing ensemble through protein-protein interactions of the constituting enzymes. Of particular interest is the observation of efficient cross-linking of CYP2E1 with CYP4A11. This enzyme plays a central role in the synthesis of vasoactive eicosanoids and its interactions with alcohol-inducible CYP2E1 may shed light on the mechanisms of alcohol-induced hypertension.

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Extensive Protein-Protein Interactions Involving UDP-glucuronosyltransferase (UGT) 2B7 in Human Liver Microsomes

Drug Metabolism and Pharmacokinetics ◽

10.2133/dmpk.dmpk-13-rg-096 ◽

2014 ◽

Vol 29 (3) ◽

pp. 259-265 ◽

Cited By ~ 13

Author(s):

Ryoichi Fujiwara ◽

Tomoo Itoh

Keyword(s):

Protein Interactions ◽

Human Liver ◽

Liver Microsomes ◽

Human Liver Microsomes ◽

Protein Protein Interactions ◽

Udp Glucuronosyltransferase

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Rotation and Protein-Protein Interactions of Cytochrome P-450 in the Inner Membrane of Adrenocortical Mitochondria

The Journal of Biochemistry ◽

10.1093/oxfordjournals.jbchem.a122439 ◽

1988 ◽

Vol 104 (2) ◽

pp. 188-191 ◽

Cited By ~ 13

Author(s):

Suguru Kawato ◽

Fumiko Mitani ◽

Tetsutaro Iizuka ◽

Yuzuru Ishimura

Keyword(s):

Protein Interactions ◽

Protein Protein Interactions ◽

Inner Membrane ◽

Cytochrome P 450

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Cytochrome P-450 and NADPH-Cytochrome c (P-450) Reductase in Reconstituted Phospholipid Vesicles as Demonstrated by Negative Staining Techniques

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100099234 ◽

1981 ◽

Vol 39 ◽

pp. 558-559

Author(s):

Jane H. Dees ◽

Linda K. Parkhill ◽

L. Dean Coe ◽

Betty Ann Germany ◽

R. T. Okita ◽

...

Keyword(s):

Cytochrome C ◽

Protein Interactions ◽

Membrane Vesicles ◽

Protein Protein Interactions ◽

Ultrastructural Studies ◽

Nadph Cytochrome ◽

Hydrophobic Tail ◽

Staining Techniques ◽

Cytochrome P 450 ◽

Structural Aspects

Artificial membrane vesicles or liposomes have become an accepted model for the study of enzyme-membrane interactions under controlled conditions.The development of reconstituted vesicle models for the microsomal mixed function oxidase enzymes has provided a system in which to study not only the biochemistry of these enzymes in a near physiologic milieu, but also structural aspects of the protein-phospholipid and protein-protein interactions. Ultrastructural studies on liposomes reconstituted with NADPH- cytochrome c (P-450) reductase (an amphipathic protein, 78,000 daltons, thought to be partially embedded in the microsomal membrane by its hydrophobic tail), and cytochrome P-450 (a hydrophobic protein, about 50,000 daltons, thought to be embedded within the lipid bilayer) either singly or together, should reveal information on whether these proteins are randomly arranged in the membrane or arranged in an ordered manner, an issue currently debated among cytochrome P-450 researchers.

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