title: Existence of metastable intermediate lysozyme conformation highlights the role of alcohols in altering protein stability.
abstract: Alcohols have a manifold effect on the conformational and thermodynamic stability of native proteins. Here, we study the effect of moderate concentrations of trifluoroethanol (TFE) on the thermal stability of hen egg-white lysozyme (HEWL), by far-UV circular dichroism and by steady-state and time-resolved photoluminescence of intrinsic tryptophans. Our results highlight that TFE affects lysozyme stability by preferential solvation of the protein molecule. Furthermore, we discovered the existence at 20% TFE of an equilibrium partially folded state of lysozyme, intermediate between the native and the unfolded state. A three-state model is therefore used to interpolate the thermal denaturation data. Our analysis explains how the stabilization of the intermediate conformation enhances the entropic contribution to unfolding, and thus decreases the unfolding temperature, while, at the same time, TFE enhances the conformational stability of the native fold at room temperature. Eventually, we challenged the ability of these intermediate structures to form supramolecular aggregates by heating experiments at different TFE concentrations.
authors.affiliation: Institute of Biophysics at Palermo (IBF), National Research Council of Italy (CNR), via U. La Malfa 153, I-90146 Palermo, Italy.
authors.forename: Michele
authors.initials: M
authors.lastname: D'Amico
authors.forename: Samuele
authors.initials: S
authors.lastname: Raccosta
authors.forename: Marco
authors.initials: M
authors.lastname: Cannas
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Martorana
authors.forename: Mauro
authors.initials: M
authors.lastname: Manno
mesh_terms: D000438:Alcohols / Q000737:chemistry*; D000818:Animals; D002645:Chickens; D002942:Circular Dichroism; D009113:Muramidase / Q000737:chemistry*; D011489:Protein Denaturation; D055550:Protein Stability; D017433:Protein Structure, Secondary; D058767:Protein Unfolding; D013696:Temperature; D014270:Trifluoroethanol / Q000737:chemistry
chemical_list: D000438:Alcohols; D014270:Trifluoroethanol; C421864:hen egg lysozyme; D009113:Muramidase