NMR studies on dynamic behavior of water molecule in aqueous denaturant solutions at 25 degrees C: Effects of guanidine hydrochloride, urea and alkylated ureas.

The spin-lattice relaxation times (T-1) of the 17(O) nucleus of a water mol ecule in guanidine hydrochloride, urea, and alkylated ureas at 25 degrees C were measured by NMR spectroscopy. Urea has no significant effect on the w ater structure. Also, guanidine hydrochloride as a stronger denaturant than urea does not perturb the water structure similar to urea. The change in t he dynamic structure (B-R) of water was proportional to the carbon number o f the alkyl chain of the alkylated urea. Compared to the results of alcohol s and tetraalkylammonium solution, it is clear that the contribution of one carbon of the alkyl chain to the water structure is independent of the typ e of solute. The B-R of urea and its alkylated substances show a good corre lation with the concentration of denaturation midpoints obtained on sperm w hale myoglobin and cytochrome c. Because the water structure is not perturb ed by urea and guanidine hydrochloride, these results indicate that the eff ect of hydrophobic hydration of denaturants becomes important for the denat uration of a protein with increasing hydrophobicity of the denaturants. (C) 2000 Elsevier Science B.V. All rights reserved.