Figure 4.

The thermal stability of monomeric (+, T_m = 49.5^oC, ΔH = 110 kcal/mol) and polymeric (○, apparent T_m = 40.0^oC, apparent ΔH = 50 kcal/mol) T21C/K124C/WT* T4 lysozyme. Monomeric and polymeric samples were thermally unfolded (18) in 20 mM glycine-HCl, 1 mM H₄EDTA, pH 3.05, with protein concentrations of 15 and 10 μM (in residue), respectively. The changes in the CD signal at 223 nm were analyzed as two-state transitions (solid curves). CD signals returned to in excess of 95% of their starting values upon cooling but approximately 10% of the polymeric sample adhered to the wall of the cuvette. The van't Hoff enthalpy at the apparent T_m for the polymeric sample is about 50% of the value expected on the basis of enthalpy changes with T_m of other T4 lysozyme mutants. The low value of ΔH for the polymeric sample likely reflects both heterogeneity inherent in a mixture as well as a loss of solubility of some of the material. The polymeric sample is less stable than the monomer. The change in ΔΔG relative to monomeric T21C/K124C/WT* is −3.5 kcal/mol with uncertainty likely ±1 kcal/mol.