Linkage of proton binding to the thermal unfolding of Sso7d from the hyperthermophilic archaebacterium Sulfolobus solfataricus

In this study the pH dependence of the thermal stability of Sso7d from Sulf olobus solfataricus is analyzed. This small globular protein of 63 residues shows a very marked dependence of thermal stability on pH: the denaturatio n temperature passes from 65.2 degrees C at pH 2.5 to 97.9 degrees C at pH 4.5. Analysis of the data points out that the binding of at least two proto ns is coupled to the thermal unfolding. By linking the proton binding to th e conformational unfolding equilibrium, a thermodynamic model, which is abl e to describe the dependence upon the solution pH of both the excess heat c apacity function and the denaturation Gibbs energy change for Sso7d, is dev eloped. The decreased stability in very acid conditions is due to the bindi ng of two protons on identical and noninteracting sites of the unfolded sta te. Actually, such sites are two carboxyl groups possessing very low pK(a) values in the native structure, probably involved in salt-bridges on the pr otein surface. (C) 1999 Elsevier Science B.V. All rights reserved.