Folding of chymotrypsin inhibitor 2. 1. Evidence for a two-state transition.


Abstract

The reversible folding and unfolding of barley chymotrypsin inhibitor 2 (CI2) appears to be a rare example in which both equilibria and kinetics are described by a two-state model. Equilibrium denaturation by guanidinium chloride and heat is completely reversible, and the data can be fitted to a simple two-state model involving only native and denatured forms. The free energy of folding in the absence of denaturant, delta GH2O, at pH 6.3, is calculated to be 7.03 +/- 0.16 and 7.18 +/- 0.43 kcal mol-1 for guanidinium chloride and thermal denaturation, respectively. Scanning microcalorimetry shows that the ratio of the van't Hoff enthalpy of denaturation to the calorimetric enthalpy of denaturation does not deviate from unity, the value observed for a two-state transition, over the pH range 2.2-3.5. The heat capacity change for denaturation is found to be 0.789 kcal mol-1 K-1. The rate of unfolding of CI2 is first order and increases exponentially with increasing guanidinium chloride concentration. Refolding, however, is complex and involves at least three well-resolved phases. The three phases result from heterogeneity of the unfolded form due to proline isomerization. The fast phase, 77% of the amplitude, corresponds to the refolding of the fraction of the protein that has all its prolines in a native trans conformation. The rate of this major phase decreases exponentially with increasing guanidinium chloride concentration. The unfolding and refolding kinetics can also be fitted to a two-state model.(ABSTRACT TRUNCATED AT 250 WORDS) Study holds ProTherm entries: 2953, 2954, 2955, 2956, 2957, 11831, 11832, 11833, 11834, 11835 Extra Details: dG was calculated at 25 degrees C

Submission Details

ID: W82SgH2Q3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:20 p.m.

Version: 1

Publication Details
Jackson SE;Fersht AR,Biochemistry (1991) Folding of chymotrypsin inhibitor 2. 1. Evidence for a two-state transition. PMID:1931967
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