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Use of selective extraction and fast chromatographic separation combined with electrophoretic methods for mapping of membrane proteins
Author(s) -
Josic Djuro,
Brown Mari Kino,
Huang Feilei,
Callanan Helen,
Ručević Marijana,
Nicoletti Alison,
Clifton James,
Hixson Douglas C.
Publication year - 2005
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200500060
Subject(s) - chromatography , chemistry , electrophoresis , membrane , extraction (chemistry) , membrane protein , urea , mass spectrometry , protein purification , ion chromatography , ion exchange , biochemistry , ion , organic chemistry
A model system for selective solubilization and fast separation of proteins from the rat liver membrane fraction and purified rat liver plasma membranes for their further proteomic analysis is presented. For selective solubilization, high‐pH solutions and a concentrated urea solution, combined with different detergents, are used. After extraction, proteins are separated by anion‐exchange chromatography or a combination of anion‐ and cation‐exchange chromatography with convective interaction monolithic supports. This separation method enables fast and effective prefractionation of membrane proteins based on their hydrophobicity and charge prior to one‐dimensional (1‐D) and 2‐D electrophoresis and mass spectrometry. By use of this sample preparation method, the less‐abundant proteins can be detected and identified.