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

As a complementary approach to genome projects, proteomic analyses have bee n set up to identify new gene products. One of the major challenges in prot eomics concerns membrane proteins, especially the minor ones. A procedure b ased on the differential extraction of membrane proteins in chloroform/meth anol mixtures, was tested on the two different chloroplast membrane systems : envelope and thylakoid membranes. Combining the use of classical sodium d odecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and mass spect rometry analyses, this procedure enabled identification of hydrophobic prot eins. The propensity of hydrophobic proteins to partition in chloroform/met hanol mixtures was directly correlated with the number of amino acid residu es/number of putative transmembrane regions (Res/TM ratio). Regardless of t he particular case of some lipid-interacting proteins, chloroform/methanol extractions allowed enrichment of hydrophobic proteins and exclusion of hyd rophilic proteins from both membrane systems, thus demonstrating the versat ility of the procedure.