Integration of a Two-Phase Partition Method into Proteomics Research on Rat Liver Plasma Membrane Proteins
- 14 February 2006
- journal article
- research article
- Published by American Chemical Society (ACS) in Journal of Proteome Research
- Vol. 5 (3), 634-642
- https://doi.org/10.1021/pr050387a
Abstract
To comprehensively identify proteins of the rat liver plasma membrane (PM), we have adopted a proteomics strategy that utilizes sucrose density centrifugation in conjunction with aqueous two-phase partition for plasma membrane isolation, followed by SDS-PAGE, mass spectrometry and bioinformatics. Western blot analysis showed that this method results in highly purified plasma membrane fractions, which is a key to successful plasma membrane proteomics. The PM proteins were separated by SDS-PAGE and digested with trypsin. Through nano-ESI−LC MS/MS analysis we identified 428 rat liver membrane proteins, of which 304 had a gene ontology (GO) annotation indicating a cellular component, and 204 (67%) of the latter were known integral membrane proteins or membrane-associated proteins. In addition to proteins known to be associated with the plasma membrane, several hypothetical proteins have also been identified. This study not only provides a tool to study plasma membrane proteins with low levels of contamination, but also provides a data set for proteins of high to moderate abundance in rat liver plasma membranes, thus allowing for more comprehensive characterization of membrane proteins and a better understanding of membrane dynamics. Keywords: rat liver • plasma membrane • plasma membrane proteins • two-phase partition • LC−MS/MSThis publication has 22 references indexed in Scilit:
- A Proteomic Analysis of Lysosomal Integral Membrane Proteins Reveals the Diverse Composition of the OrganelleMolecular & Cellular Proteomics, 2005
- Integration of Jacobson's Pellicle Method into Proteomic Strategies for Plasma Membrane ProteinsJournal of Proteome Research, 2004
- A Proteomic Characterization of the Plasma Membrane of Human Epidermis by High-Throughput Mass SpectrometryJournal of Investigative Dermatology, 2004
- Analysis, statistical validation and dissemination of large-scale proteomics datasets generated by tandem MSDrug Discovery Today, 2004
- Protein Identification by Mass SpectrometryMolecular & Cellular Proteomics, 2004
- Proteomic analysis of the mammalian nuclear pore complexThe Journal of cell biology, 2002
- Predicting transmembrane protein topology with a hidden markov model: application to complete genomesJournal of Molecular Biology, 2001
- Probability-based protein identification by searching sequence databases using mass spectrometry dataElectrophoresis, 1999
- Two-dimensional mapping of the endogenous proteins of the rat hepatocyte Golgi complex cleared of proteins in transitElectrophoresis, 1997
- A simple method for displaying the hydropathic character of a proteinJournal of Molecular Biology, 1982