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J Proteome Res. 2013 Oct 4;12(10):4402-13. doi: 10.1021/pr4004213. Epub 2013 Sep 26.

Targeted quantitative proteomics for the analysis of 14 UGT1As and -2Bs in human liver using NanoUPLC-MS/MS with selected reaction monitoring.

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Division of Molecular Pharmaceutics, Eshelman School of Pharmacy, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina 27599, United States.


Targeted quantitative proteomics using heavy isotope dilution techniques is increasingly being utilized to quantify proteins, including UGT enzymes, in biological matrices. Here we present a multiplexed method using nanoLC-MS/MS and multiple reaction monitoring (MRM) to quantify 14 UGT1As and UGT2Bs in liver matrices. Where feasible, we employ two or more proteotypic peptides per protein, with only four proteins quantified with only one proteotypic peptide. We apply the method to analysis of a library of 60 human liver microsome (HLM) and matching S9 samples. Ten of the UGT isoforms could be detected in liver, and the expression of each was consistent with mRNA expression reported in the literature. UGT2B17 was unusual in that ∼30% of liver microsomes had no or little (<0.5 pmol/mg protein) content, consistent with a known common polymorphism. Liver S9 UGT concentrations were approximately 10-15% those of microsomes. The method was robust, precise, and reproducible and provides novel UGT expression data in human liver that will benefit rational approaches to evaluate metabolism in drug development.

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