Stabilization And Destabilization By Co-Solvents (Molecular Biology)

The effect of a co-solvent on the stability of a protein (or other macromolecule) is determined strictly by the difference between the interactions of the co-solvent with the protein in the unfolded and native states (see Protein Stability). The essential interactions that must be known are preferential binding (or preferential hydation) and the transfer free energy, ie, free energy of interaction with (binding to) the protein of the structure stabilizer or destabilizer (1).

Let the unfolding (stability) process be represented by a simple equilibrium between the native N and denatured D states, and let this equilibrium be affected by a ligand (stabilizing or destabilizing co-solvent):

The effect of the co-solvent can be expressed then relative to either of two reference states:

1. In the first reference state the given co-solvent concentration is given. The question is: Does the co-solvent at this concentration stabilize the protein relative to an infinitesimally smaller concentration? The effect the co-solvent has then is expressed by the Wyman linkage relation:

where nD and nN stand for the preferential binding of the co-solvent to the unfolded and native protein (a L is the thermodynamic activity of the co-solvent, which frequently may be approximated by the concentration). Therefore, the slope of a log-log plot of the equilibrium constant versus the ligand concentration gives the difference in preferential binding between the two end states. Conversely, knowledge of and nN indicates whether a ligand (co-solvent) will be a stabilizer or destabilizer of the structure: Greater equilibrium dialysis binding to the unfolded form enhances the unfolding reaction; greater binding to the native form results in stabilization of the native structure.

(2) In the second reference state, the effect is measured relative to water as solvent. It is expressed by the difference between the binding free energies to the two end states:

where DG °L is the standard free energy of the unfolding equilibrium at ligand concentration L and DG 0w is that in water (this is related to the equilibrium constant by DG0 = -2.303RT logK). Dmpr r is the transfer free energy of the protein from water (dilute buffer) to the solvent of the given composition in the denatured D and native N states; it, in fact, is the free energy of binding of the ligand (co-solvent) to the protein (DGb = Dmpr,tr) in its two states.