Limits of Proteomics: Protein Solubilisation Issues

Thierry Rabilloud, CEA‐Grenoble, Grenoble, France

Published online: March 2012

DOI: 10.1002/9780470015902.a0006201.pub2

Because of their tremendous chemical heterogeneity, many proteins, such as membrane proteins, are poorly soluble in aqueous solvents. This precludes their analysis by classical proteomics techniques, whether these techniques are based on protein separation (e.g. two‐dimensional gel‐based proteomics) or on peptide separation after in‐solution digestion. For techniques based on peptide separation, the issue is to couple on the one side a complete solubilisation of proteins, a good efficiency of the protease used to produce the peptides, and on the other side minimal interference with the techniques used to separate the peptides. For techniques based on protein separation, the issue is to solubilise the proteins and to keep them soluble under the conditions used for the protein separation. For both type of technologies, combinations of detergents and chaotropes often provide the most performing solution to the protein solubilisation problem, although complete solubilisation can never be guaranteed.

Key Concepts:

  • Proteins must be extracted from their cellular context into an aqueous‐based solvent for their analysis.
  • Surfactants/detergents provide the adequate medium to mimic a lipid‐like environment in water.
  • The structural variety of detergents provides the various solubilisation performances that can be adapted to many proteomics methods.
  • Chaotropes are often used in conjunction with detergents to improve protein/peptide solubility.

Keywords: proteomics; detergents; membrane; chaotropes; solubilisation

Figure 1. The successive steps of protein solubilisation by detergents: (a) The structure of a biological membrane is schematised. The orange layer symbolises the lipids, the transmembrane proteins are symbolised in dark blue and the accessory subunits in various colors. (b) The first step of solubilisation, where strongly bound lipids stick to the membrane proteins and are covered by a detergent layer that makes the whole complex soluble in water. (c) The detergent molecules have completely replaced the lipids, but the proteins still keep their native structure. (d) The proteins are denatured by the interaction with the detergent and the whole polypeptide chain is embedded in a protein–detergent complex.
Figure 2. Chemical structures of detergents and chaotropes. (a–c) Ionic detergents: (a) soap (anionic detergent); (b) dodecyl trimethylammonium bromide (cationic detergent) and (c) sodium dodecyl sulfate (SDS). All these detergents are strongly denaturing to proteins but are excellent protein solubilisers. (d and e) Nonionic detergents: (d) Brij 30 (dodecyl tetraethylene glycol ether) and (e) dodecyl maltoside. Nonionic detergents are usually very mild (and thus rather poor solubilisers alone). (f and g) Zwitterionic detergents. These detergents are electrically neutral because they bear in a single molecule an equal number of negative and positive charges: (f) sulfobetaine 3–12 (SB 3–12) and (g) CHAPS. Zwitterionic detergents vary widely in their properties. CHAPS is nondenaturing but also a weak solubiliser, whereas SB 3–12 is intermediate between neutral and ionic detergents in both its solubilising and denaturing effects. Note that detergents (a–f) bear the same hydrophobic part (h). The structure of urea, the most widely used chaotrope. Note the absence of any hydrophobic part in the urea molecule, opposite to all detergents molecules.
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Related Sub-Topics:

Basic Cell Biology, Protein, RNA and DNA | Techniques in Cell Biology | Membrane Proteins | Protein Structure | Sample Preparation in Structural Biology | Biochemical and Biophysical Techniques | Proteins: Structure, Function, Metabolism
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Article Title: Limits of Proteomics: Protein Solubilisation Issues
 
How to Cite close
Rabilloud, Thierry(Mar 2012) Limits of Proteomics: Protein Solubilisation Issues. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0006201.pub2]