Determination of enzyme global thermostability from equilibrium and kinetic analysis of heat inactivation

Abstract An equilibrium-kinetic description for the two-stage irreversible thermoinactivation of enzymes (N ⇔ D ⇒ I) is examined by using chymotrypsin as a model. The parameter ΔG for enzyme unfolding (N ⇔ D) and activation free energy for the D ⇒ I reaction ( ΔG # i ) were summed to provide an index of enzyme global thermostability ( ΔG # ; ΔG # = ΔG + ΔG # i ). There was good agreement between calculated and experimental global thermostability (i.e. enzyme stability with respect to reversible and irreversible thermoinactivation reaction steps) at 0–60°C. The results indicate that enzyme global thermostability may be markedly dependent on the rate of enzyme folding.