Effect of polyol osmolytes on DeltaG(D), the Gibbs energy of stabilisation of proteins at different pH values.
Thermal denaturation curves of lysozyme and ribonuclease-A were determined by measuring their far-UV circular dichroism (CD) spectra in the presence of different concentrations of five polyols (sorbitol, glycerol, mannitol, xylitol and adonitol) at various pH values in the range 7.0--1.9. The denaturation curve at each polyol concentration and pH was analysed to obtain values of T(m) (midpoint of denaturation) and DeltaH(m) (enthalpy change at T(m)), and these DeltaH(m) and T(m) values obtained at different pH values were used to obtain DeltaC(p) (constant-pressure heat capacity change) at each polyol concentration. Using values of DeltaH(m), T(m) and DeltaC(p) in the Gibbs-Helmholtz equation, DeltaG(D) degrees (Gibbs energy change at 25 degrees C) was determined at a given pH and polyol concentration. Main conclusions of this study are that polyols have no significant effect on DeltaG(D) degrees at pH 7.0, and they stabilise proteins in terms of DeltaG(D) degrees against heat denaturation at lower pH values. Other conclusions of this study are: (i) T(m) at each pH increases with increasing polyol concentration, (ii) DeltaH(m) remains, within experimental error, unperturbed in the presence of polyols, and (iii) DeltaC(p) depends on polyol concentration. Furthermore, measurements of the far- and near-UV CD spectra suggested that secondary and tertiary structures of both proteins in their native and denatured states are not perturbed on the addition of polyols.
Haque
Inamul
I
Department of Biosciences, Jamia Millia Islamia, Jamia Nagar, New Delhi-110 025, India.
Singh
Rajendrakumar
R
Moosavi-Movahedi
Ali Akbar
AA
Ahmad
Faizan
F
D004734:Energy Metabolism / Q000187:drug effects; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D011108:Polymers / Q000494:pharmacology*; D011489:Protein Denaturation / Q000187:drug effects*; D012259:Ribonuclease, Pancreatic / Q000737:chemistry* / Q000187:drug effects; D013816:Thermodynamics
D011108:Polymers; C024617:polyol; D012259:Ribonuclease, Pancreatic