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I'm struggling to find peer reviewed literature that explains the effect of changing the pH and the salt concentration on protein/protein complexes in solution. What effect does the pH and the salt concentration have and what is the physical basis behind this?

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I submitted an edit to your question, it was not entirely clear to me if you ask about proteins alone or protein complexes and interactions. If I got it wrong please just edit your post again. –  Mad Scientist Dec 29 '11 at 21:35
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1 Answer 1

up vote 13 down vote accepted

The formation of protein complexes or aggregates in aqueous buffers is determined by a number of factors: physical properties of the protein itself, pH, temperature, type and concentration of the used cosolvent (salt). Solutes are often roughly divided by type into chaotropes ('disorder-making'), which destabilise protein structures and kosmotropes ('order-making'), which stabilize them. [1, 2]

Chaotropic salts interfere with intramolecular interactions mediated by non-covalent forces such as hydrogen bonds, van der Waals forces, and hydrophobic interactions, which, at high cosolvent concentrations, results in protein denaturation.

Kosmotropic salts, on the other hand, cause water molecules to favorably interact, which also stabilizes intermolecular interactions in proteins. The salt molecules readily interact with water from the protein's hydratation shell and remove it from the protein surface, which produces thermodynamically unfavourable interactions that are reduced when proteins associate to form complexes. With increase in salt concentration the protein precipitation (salting out) increases.

Both cations and anions have been ranked separately by their capacity to precipitate proteins, to form a Hofmeister series [3, 4] (first set in 1888 but still hotly debated), e.g.:

SO42− > H2PO4 > CH3COO > Cl > Br

At large salt concentrations protein solubility is given by the empirical Cohn equation [6]:

lnS = α − βc

where S is the protein solubility, c is the salt's ionic strength, α and β are empirical constants characteristic of particular salt.

Salting-out agents are very widely used in protein purification (to concentrate proteins eg. with ammonium sulfate), chromatography or crystalization.

The influence of pH on protein-protein interactions in solution works through altering of the electrostatic properties of protein surfaces. At pH equal to the protein's isoelectric point (pI), where its net charge is neutral, charge repulsions of similar molecules are relatively low and many proteins will aggregate. Very low and very high pH will case proteins to denature; during digestion, for instance, proteins are in extremely low and then extremly high pH that exposes their backbones for enzymatic degradation.

For more information, please refer to:

  1. Martin Chaplin, Kosmotropes and Chaotropes, http://www.lsbu.ac.uk/water/kosmos.html

  2. Zangi R. Can salting-in/salting-out ions be classified as chaotropes/kosmotropes? J Phys Chem B. 2010 Jan 14;114(1):643-50.

  3. Pace CN, Treviño S, Prabhakaran E, Scholtz JM. Protein structure, stability and solubility in water and other solvents. Philos Trans R Soc Lond B Biol Sci. 2004 Aug 29;359(1448):1225-34; discussion 1234-5.

  4. Shimizu S, McLaren WM, Matubayasi N. The Hofmeister series and protein-salt interactions. J Chem Phys. 2006 Jun 21;124(23):234905.

  5. Zhang Y, Cremer PS. Interactions between macromolecules and ions: The Hofmeister series. Curr Opin Chem Biol. 2006 Dec;10(6):658-63. Epub 2006 Oct 10.

  6. Ruckenstein E, Shulgin IL. Effect of salts and organic additives on the solubility of proteins in aqueous solutions. Adv Colloid Interface Sci. 2006 Nov 16;123-126:97-103. Epub 2006 Jun 30.

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