Thermal stability of calf skin collagen type I in salt solutions

The thermal stability of acid-soluble collagen type I from calf skin in salt solutions is studied by high-sensitivity differential scanning calorimetry. Three concentration ranges have been clearly distinguished in the dependence of collagen thermal stability on ion concentration. At concentrations below 20 mM, all studied salts reduce the temperature of collagen denaturation with a factor of about 0.2 degree C per 1 mM. This effect is attributed to screening of electrostatic interactions leading to collagen stabilisation. At higher concentrations, roughly in the range 20-500 mM, the different salts either slightly stabilise or further destabilise the collagen molecule in salt-specific way that correlates with their position in the lyotropic series. The effect of anions is dominating and follows the order H2PO4- > or = SO4(2-) > Cl- > SCN-, with sign inversion at about SO4(2-). This effect, generally known as the Hofmeister effect, is associated with indirect protein-salt interactions exerted via competition for water molecules between ions and the protein surface. At still higher salt concentrations (onset concentrations between 200 and 800 mM for the different salts), the temperature of collagen denaturation and solution opacity markedly increase for all studied salts due to protein salting out and aggregation. The ability of salts to salt out collagen also correlates with their position in the lyotropic series and increases for chaotropic ions. The SO4(2-) anions interact specifically with collagen - they induce splitting of the protein denaturation peak into two components in the range 100-150 mM Na2SO4 and 300-750 mM Li2SO4. The variations of the collagen denaturation enthalpy at low and intermediate salt concentrations are consistent with a weak linear increase of the enthalpy with denaturation temperature. Its derivative, d(delta H)/dT, is approximately equal to the independently measured difference in the heat capacities of the denatured and native states, delta Cp = Cp(D) - Cp(N) approximately 0.1 cal.g-1 K-1.