Conformational stability of the covalent complex between elastase and alpha 1-proteinase inhibitor.

The equilibrium unfolding-refolding process of the elastase-alpha 1-proteinase inhibitor complex, induced by guanidinium chloride, was followed by spectroscopic methods. A reversible transition with a midpoint at 2.04 +/- 0.04 M guanidinium chloride was observed by fluorescence. This transition was attributed to elastase on the basis of circular dichroism and uv absorption difference data obtained for the covalent complex and for the free proteins. The conformational stability of elastase in the complex was analyzed considering the approximation of a two-state transition. The free energy of denaturation delta GH2O was 4.2 kcal.mol-1 for complexed elastase compared to 10.5 kcal.mol-1 for the free enzyme. Such a decrease in the stability of elastase suggests that, after forming the covalent complex with the inhibitor, the enzyme undergoes not only the expected local modifications of the active site, but also an extensive structural reorganization.