SUCROSE REDUCES THE EFFICIENCY OF PROTEIN DENATURATION BY A CHAOTROPIC AGENT

Sugars and polyols are used to stabilize proteins. The degree of stabi lization conferred on a model protein by sucrose was calculated in ter ms of the free energy of folding. Phosphoglycerate kinase (PGK) was de natured by guanidine hydrochloride (GuHCl) in different sucrose concen trations. The linear extrapolation method [1,2] was used to calculate the free energy of folding in the absence of denaturant. Although sucr ose increased the concentration of GuHCl required to unfold the protei n, the free energy of folding in water was unchanged. In order to prob e the nature of the stabilizing effect of sucrose, an FT-Raman spectro scopic study of denaturant-polyol systems was undertaken. Investigatio ns of interactions between GuHCl, urea or formamide and polyhydric com pounds, revealed no evidence for hydrogen bonding or dipole-dipole ass ociations. Polyhydric compounds caused minor changes in denaturant spe ctra although the converse was not observed. The structure of deuterat ed water changed on addition of denaturants. For non-ionic denaturants , addition of polyhydric solutes countered this change in water struct ure. Thus polyhydric compounds oppose the effect of denaturants on wat er structure. The observed increase in GuHCl concentration required to unfold PGK in the presence of sucrose may be attributed to this prope rty of sucrose.