Extraction of Escherichia coli proteins with organic solvents prior to two-dimensional electrophoresis

Mark P. Molloy, Ben R. Herbert, Keith L. Williams*, Andrew A. Gooley

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review


Compared to soluble proteins, hydrophobic proteins, in particular membrane proteins, are an underrepresented protein species on two-dimensional (2-D) gels. One possibility is that many hydrophobic proteins are simply not extracted from the sample prior to 2-D gel separation. We attempted to isolate hydrophobic proteins from Escherichia coli by extracting with organic solvents, then reconstituting the extracted proteins in highly solubilising sample solution amenable to 2-D electrophoresis using immobilized pH gradients (IPGs). This was conducted by an extraction with a mixture of chloroform and methanol, followed by solubilisation using a combination of urea, thiourea, sulfobe-taine detergents and tributyl phosphine. Peptide mass fingerprinting assisted in the identification of 13 proteins, 8 of which have not previously been reported on 2-D gels. Five of these new proteins possess a positive hydropathy plot. These results suggest that organic solvent extractions may be useful for selectively isolating some proteins that have previously been missing from proteome maps.

Original languageEnglish
Title of host publicationFrom genome to proteome
Subtitle of host publicationadvances in the practice & application of proteomics
EditorsMichael Dunn
Place of PublicationWeinheim, Germany; New York
PublisherWiley-Blackwell, Wiley
Number of pages4
ISBN (Electronic)9783527613489
ISBN (Print)9783527301546, 9783527613496, 3527301542
Publication statusPublished - 26 Dec 2007


  • Extraction
  • Organic solvent
  • Proteome
  • Solubility
  • Two-dimensional electrophoresis


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