Influence of glycerol on the cooling effect of pair hydrophobicity in water: relevance to proteins' stabilization at low temperature.

Abstract

Glycerol, as a cosolvent of water, stabilizes proteins under extreme conditions (both at high and low temperatures). However, the mechanism of stabilization of proteins by glycerol at low temperature is still elusive. Because the decrease of hydrophobic interactions at a lower temperature is one of the crucial factors for the cold denaturation, we ask here whether glycerol protects the hydrophobic interactions upon cooling and thereby acts against cold denaturation. Here, we have performed potential of mean force (PMF) calculations, using the umbrella sampling technique, between a pair of methane hydrophobic solute molecules either in pure water or in binary mixtures of water and glycerol for two different compositions and each of them at four different temperatures. We have found that glycerol increases the pair hydrophobic interaction at all the temperatures studied and that the enhancement is more prominent at the lower temperatures studied here. Decomposition of the PMF into the enthalpic and the entropic components and detailed molecular structural analyses give insight into the above observation. We have found that the enhancement of the hydrophobic interaction with increasing glycerol concentration occurs primarily due to the strengthening of the glycerol-water interaction near the associated methane solute molecule pair and the tetrahedral ordering of the H-bonding network being made uniform around the solute by the added glycerol molecules. These results indirectly justify the efficacy of glycerol for the preservation of proteins against cold denaturation at low temperature.