Two-dimensional electrophoretic separation of subproteomes in adult rat hippocampal subregions after membrane protein enrichment with sequential extraction

Abstract

Abstract: Objective To establish a method for analysis of the subproteomes following membrane protein enrichment, so as to investigate the differences in cytosolic and membrane protein/peptide contents among different hippocampal subregions (CA1, CA3 and dentate gyrus) of rats.Methods The hippocampal subregions were isolated by microdissection, and after membrane protein enrichment with protein solubility-based sequential sample extraction, subproteome profiling was accomplished using modified two-dimensional electrophoresis and silver staining method compatible with high-performance mass spectrometry, which displayed cytosolic or membrane proteins/peptides separately. The efficacy of the electrophoresis was evaluated by image analyzing software.Results The membrane proteins were enriched by 8 folds, and around 30% more spots were shown on the integrated dentate gyrus map than on the conventional map obtained by one-step protein extraction. Such improvement could also be seen in protein mapping of CA1 and CA3 regions.Conclusion When analyzing hippocampal subregions sequential extraction and two-dimensional electrophoresis more effectively separate and display enriched membrane protein/peptides, which may play important roles in such cellular events as signal transduction.

CLC Number:

R338.2

CHEN Liang¹, YANG Jian-ming¹, LI Xiao-ming¹, LI Xiao-wen¹, GAO Tian-ming^1,2. Two-dimensional electrophoretic separation of subproteomes in adult rat hippocampal subregions after membrane protein enrichment with sequential extraction[J]. Journal of Southern Medical University, 2004, 24(11): 1217-1222,1226.

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