Effect of monomeric and oligomeric sugar osmolytes on DeltaGD, the Gibbs energy of stabilization of the protein at different pH values: is the sum effect of monosaccharide individually additive in a mixture?
Thermal denaturation curves of ribonuclease-A were measured by monitoring changes in the far-UV circular dichroism (CD) spectra in the presence of different concentrations of six sugars (glucose, fructose, galactose, sucrose, raffinose and stachyose) and mixture of monosaccharide constituents of each oligosaccharide at various pH values in the range of 6.0-2.0. These measurements gave values of T(m) (midpoint of denaturation), DeltaH(m) (enthalpy change at T(m)), DeltaC(p) (constant-pressure heat capacity change) under a given solvent condition. Using these values of DeltaH(m), T(m) and DeltaC(p) in appropriate thermodynamic relations, thermodynamic parameters at 25 degrees C, namely, DeltaG(D)(o) (Gibbs energy change), DeltaH(D)(o) (enthalpy change), and DeltaS(D)(o) (entropy change) were determined at a given pH and concentration of each sugar (including its mixture of monosaccharide constituents). Our main conclusions are: (i) each sugar stabilizes the protein in terms of T(m) and DeltaG(D)(o), and this stabilization is under enthalpic control, (ii) the protein stabilization by the oligosaccharide is significantly less than that by the equimolar concentration of the constituent monosaccharides, and (iii) the stabilization by monosaccharides in a mixture is fully additive. Furthermore, measurements of the far- and near-UV CD spectra suggested that secondary and tertiary structures of protein in their native and denatured states are not perturbed on the addition of sugars.
Poddar
Nitesh Kumar
NK
Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi-110025, India.
Ansari
Z A
ZA
Singh
R K Brojen
RK
Moosavi-Movahedi
A A
AA
Ahmad
Faizan
F
D002942:Circular Dichroism; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D009005:Monosaccharides / Q000737:chemistry*; D009844:Oligosaccharides / Q000737:chemistry*; D009994:Osmolar Concentration; D011487:Protein Conformation; D011489:Protein Denaturation; D012259:Ribonuclease, Pancreatic / Q000737:chemistry*; D013816:Thermodynamics
D009005:Monosaccharides; D009844:Oligosaccharides; D012259:Ribonuclease, Pancreatic