Transfer free energies and average static accessibilities for ribonuclease A in guanidinium hydrochloride and urea solutions

Abstract

Isopiestic vapor pressure measurements were used to obtain free energies of transfer [.DELTA.Gtr] of RNase A [EC 3.1.4.] from dilute buffer to solutions of either urea or guanidine hydrochloride (GdnHCl) over a wide cosolute concentration range. .DELTA.Gtr vary monotonically from 0 to -8 kcal/mol in 8 M urea and to -18 kcal/mol in 6 M GdnHCl. These values are not large in relation to .DELTA.Gtr of constituent groups of the protein from water to cosolute solutions of the same concentration. The assumption is made that the magnitude of .DELTA.Gtr of the protein is governed by the average static accessibility of its constituent groups to the solution. .DELTA.Gtr to different cosolute concentrations of a hypothetical 100% accessible RNase A were determined using literature values of the .DELTA.Gtr of constituent groups and the amino acid composition. The ratio of the experimentally determined .DELTA.Gtr of the 100% accessible protein gave 11% accessible surface area for the native protein in 1 M GdnHCl or 2 M urea. Additional considerations led to a value of 36% for the accessible surface area of the denatured protein in 6 M GdnHCl or 8 M urea.