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Organic solvent extraction as a versatile procedure to identify hydrophobic chloroplast membrane proteins.

Electrophoresis (2000-11-18)
M Ferro, D Seigneurin-Berny, N Rolland, A Chapel, D Salvi, J Garin, J Joyard
ABSTRACT

As a complementary approach to genome projects, proteomic analyses have been set up to identify new gene products. One of the major challenges in proteomics concerns membrane proteins, especially the minor ones. A procedure based on the differential extraction of membrane proteins in chloroform/methanol mixtures, was tested on the two different chloroplast membrane systems: envolope and thylakoid membranes. Combining the use of classical sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry analyses, this procedure enabled identification of hydrophobic proteins. The propensity of hydrophobic proteins to partition in chloroform/methanol mixtures was directly correlated with the number of amino acid residues/number of putative transmembrane regions (Res/TM ratio). Regardless of the particular case of some lipid-interacting proteins, chloroform/methanol extractions allowed enrichment of hydrophobic proteins and exclusion of hydrophilic proteins from both membrane systems, thus demonstrating the versatility of the procedure.

MATERIALS
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Product Description

Sigma-Aldrich
Plant Fractionated Protein Extraction Kit, Suitable for any plant species or tissue