title: Simultaneous determination of structural and thermodynamic effects of carbohydrate solutes on the thermal stability of ribonuclease A.
abstract: This communication describes a new technique for the study of the effects of carbohydrates on the thermal stability of proteins. This approach combines capillary electrophoresis (CE) and protein charge ladders, collections of proteins that differ incrementally in number of chemically modified charged groups, to provide information on both the thermodynamics (i.e., the free energy, DeltaGN-D, of denaturation), and structural changes (i.e., the effective hydrodynamic radius, RH, of proteins in both the native and denatured states) associated with stability. This information, obtained in a single set of electrophoresis experiments, allows a simple microscopic interpretation of the effects of carbohydrate solutes on protein stability. We use this technique to show that the stabilization of ribonuclease A at pH 8.4 by sucrose and fructose can be explained entirely by the contribution these solutes make to the entropy of formation of the protein-solution interface. There is no need, in this case, to refer to quasichemical concepts such as preferential hydration, binding, or exchange of solutes with water at specific sites on the protein to account for the stabilizing effects observed.
authors.affiliation: Department of Chemical Engineering, Princeton University, Princeton, New Jersey 08544, USA.
authors.forename: Thomas F
authors.initials: TF
authors.lastname: O'Connor
authors.forename: Pablo G
authors.initials: PG
authors.lastname: Debenedetti
authors.forename: Jeffrey D
authors.initials: JD
authors.lastname: Carbeck
mesh_terms: D004795:Enzyme Stability; D005632:Fructose / Q000737:chemistry*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D009994:Osmolar Concentration; D011489:Protein Denaturation; D012259:Ribonuclease, Pancreatic / Q000737:chemistry*; D012996:Solutions; D013395:Sucrose / Q000737:chemistry*; D013816:Thermodynamics
chemical_list: D012996:Solutions; D005632:Fructose; D013395:Sucrose; D012259:Ribonuclease, Pancreatic