Effect of the central disulfide bond on the unfolding behavior of elongation factor Ts homodimer from Thermus thermophilus

Functionally active elongation factor Ts (EF-Ts) from Thermus thermophilus forms a homodimer. The dimerization interface of EF-Ts is composed of two a ntiparallel P-sheets that can be connected by an intermolecular disulfide b ond. The stability of EF-Ts from T. thermophilus in the presence and absenc e of the intermolecular disulfide bond was studied by differential scanning calorimetry and circular dichroism. The ratio of the van't Hoff and calori metric enthalpies, Delta (vH)/DeltaH(cal), indicates that EF-Ts undergoes t hermal unfolding as a dimer independently of the presence or absence of the disulfide bond. This can be concluded from (1) the presence of residual se condary structure above the thermal transition temperature, (2) the absence of concentration dependence, which would be expected for dissociation of t he dimer prior to unfolding of the monomers, and (3) a relatively low heat capacity change (AC,) upon unfolding. The retained dimeric structure of the thermally denatured state allowed for the determination of the effect of t he intermolecular disulfide bond on the conformational stability of EF-Ts, which is Delta DeltaG((S-S,SH HS)) = 10.5 kJ/mol per monomer at 72.5 degree sC. The possible physiological implications of the dimeric EF-Ts structure and of the intersubunit disulfide bond for the extreme conformational stabi lity of proteins in thermophiles are discussed.