title: Targeted label-free approach for quantification of epoxide hydrolase and glutathione transferases in microsomes.
abstract: The aim of this study was to investigate the expression and organ distribution of cytochrome P450 (CYP450) enzymes, microsomal epoxide hydrolase (MEH), and microsomal glutathione-S-transferase (MGST 1, 2, 3) in human liver, lung, intestinal, and kidney microsomes by targeted peptide-based quantification using nano liquid chromatography-tandem multiple reaction monitoring (nano LC-MRM). Applying this method, we analyzed 16 human liver microsomes and pooled lung, kidney, and intestine microsomes. Nine of the CYP450s (CYP1A2, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, 3A4, 3A5) could be quantified in liver. Except for CYP3A4 and 3A5 existing in intestine, other CYP450s had little content (<0.1 pmol/mg protein) in extrahepatic tissues. MEH and MGSTs could be quantified both in hepatic and in extrahepatic tissues. The highest concentrations of MEH and MGST 1, 2 were found in liver; conversely MGST 3 was abundant in human kidney and intestine compared to liver. The targeted proteomics assay described here can be broadly and efficiently utilized as a tool for investigating the targeted proteins. The method also provides novel CYP450s, MEH, and MGSTs expression data in human hepatic and extrahepatic tissues that will benefit rational approaches to evaluate metabolism in drug development.
authors.affiliation: Institute of Resource Biology and Biotechnology, Department of biotechnology, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan 430074, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Song
authors.affiliation: Institute of Resource Biology and Biotechnology, Department of biotechnology, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan 430074, China. Electronic address: yulongjiang@mail.hust.edu.cn.
authors.forename: Longjiang
authors.initials: L
authors.lastname: Yu
authors.affiliation: Hubei Province Key Laboratory of Biotechnology of Chinese Traditional Medicine, Hubei University, Wuhan 430062, China. Electronic address: Peng_Zhihong@hotmail.com.
authors.forename: Zhihong
authors.initials: Z
authors.lastname: Peng
mesh_terms: D000595:Amino Acid Sequence; D002853:Chromatography, Liquid / Q000379:methods; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis*; D057075:Enzyme Assays / Q000379:methods; D004851:Epoxide Hydrolases / Q000032:analysis*; D005982:Glutathione Transferase / Q000032:analysis*; D006801:Humans; D007422:Intestines / Q000201:enzymology; D007668:Kidney / Q000201:enzymology; D008168:Lung / Q000201:enzymology; D008862:Microsomes, Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D053719:Tandem Mass Spectrometry / Q000379:methods
chemical_list: D003577:Cytochrome P-450 Enzyme System; D005982:Glutathione Transferase; D004851:Epoxide Hydrolases
keywords: Cytochrome P450 enzymes; Microsomal epoxide hydrolase; Microsomal glutathione-S-transferases; Microsomes; Nano liquid chromatography–tandem multiple reaction monitoring