To solubilise proteins you need to break proteins apart (see guide to protein-protein interactions for details on the forces that keep proteins together) and keep them apart. 

 

Physical disruption: These are your typical lysis methods including bead beating, freeze fracture, sonication etc. In BioMS our preferred method of disruption is focussed ultrasonication using Covaris AFA systems. The reason for this is that the systems are highly effective in introducing disruptive energy to a system without substantially increasing heat load. They can also be used on any sample type and process up to 96 samples simultaneously to ensure that all samples within a batch are treated equally and variability is minimised.

 

Chemical disruption: The two main non-covalent interactions that are important to protein solubility are electrostatic interactions (a positive charge attracted to a negative charge) and hydrophobic interactions (water pushing together hydrophobic areas of protein). Electrostatic interactions are relatively easy to break by increasing the ionic strength of the buffer solution (e.g. adding salt), by removing the charge (e.g. neutralising weak acids by dropping the pH) or by coating the protein in a single charge (e.g. adding a charged detergent like SDS). Hydrophobic interactions can be broken by either reducing the structure of water (e.g. adding organic solvents) or by making the hydrophobic regions more hydrophilic (e.g. by coating them in detergent). There are some compounds called chaotropes (e.g. urea and guanidine) that can also help solubilise proteins but their precise mechanisms have been subject to debate.

 

When assessing a solubilising agent it is important to know the types of interactions that the agent disrupts e.g. adding salt disrupts electrostatic interactions but amplifies hydrophobic interactions, urea and organic solvents disrupt hydrophobic interactions but not electrostatic. When a solubilising agent disrupts the interactions that make a protein fold but do not prevent new interactions forming then precipitation can occur e.g. adding organic solvent to a protein solution reduced the hydrogen bonding in the water allowing hydrophobic areas of a protein become exposed that can either bind or entangle with other proteins leading to aggregation and precipitation.

 

Proteins that are integrated into lipid bilayers are particularly difficult to solubilise as not only are they hydrophobic but they also require the lipid bilayer to be disrupted before they release. In these instances special detergents that mimic different types of lipid may be required (e.g. bile salt type detergents) and the combinations required may only be determined experimentally.

For an excellent overview of the issues around protein solubility click here