生成人类肝脏细胞色素P450的一维蛋白质组图谱。

Producing a one-dimensional proteomic map for human liver cytochromes p450.

作者信息

Petushkova Natalia A, Lisitsa Andrey V

机构信息

Institute of Biomedical Chemistry, Russian Academy of Medical Sciences, Moscow, Russia.

出版信息

Methods Mol Biol. 2012;909:63-82. doi: 10.1007/978-1-61779-959-4_5.

DOI:10.1007/978-1-61779-959-4_5

PMID:22903709

Abstract

In this chapter we explore the inducible cytochrome P450 (CYP) forms as an example of membrane proteins analysis that relies on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) fractionation with subsequent mass spectrometric (MS) identification. The approach involves cutting an SDS-PAGE gel lane into thin slices and identifying proteins in each slice by MS with the aim of obtaining detailed information on proteins of interest. A one-dimensional proteomic map showing the distribution of selected CYP isoforms across 40 slices was constructed using mass spectra obtained from each slice. Our protocol proved to be efficient enough to obtain a comprehensive profile of drug-metabolizing enzymes in the human liver. In addition to human tissues, the approach described should be applicable to the characterization of membrane proteins in other eukaryotic species.

摘要

在本章中，我们以可诱导细胞色素P450（CYP）形式为例，探讨基于十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）分级分离及后续质谱（MS）鉴定的膜蛋白分析方法。该方法包括将SDS-PAGE凝胶泳道切成薄片，并通过质谱鉴定每片中的蛋白质，目的是获取有关目标蛋白质的详细信息。利用从每片中获得的质谱构建了一张一维蛋白质组图谱，展示了选定的CYP同工型在40个切片中的分布情况。我们的实验方案被证明足够有效，能够获得人类肝脏中药物代谢酶的全面概况。除了人体组织外，所述方法应适用于其他真核生物物种膜蛋白的表征。