Studies On Displacement Adsorption Enthalpies And Conformational Change Of Denatured Bovine Serum Albumin On Hydrophobic Surface

Bovine Serum Albumin (BSA) was selected as a research object and the microcalorimetric measurement of displacement adsorption enthalpiesΔH of denatured BSA adsorbed onto a moderately hydrophobic surface (the end-group of polyethylene glycol PEG-600) from solutions (various concentrations of ammonium sulfate ((NH₄)₂SO₄) and guanidine hydrochloride (GuHCl), 0.05 mol·L-1 KH2PO4, pH 7.0) at 25℃were carried out. The thermodynamic functions and their fractions were calculated based on the stoichiometric displacement theory for adsorption (SDT-A) and its thermodynamics in combination with adsorption isotherms. The contents of secondary structure elements and thermal stability of adsorbed BSA molecules were determined by FTIR and DSC respectively. The rule of the effect of GuHCl and (NH₄)₂SO₄ on the conformational change were explored to provide theory evidence for in search of the optimal refolding path of denatured protein on the hydrophobic surface.The adsorption isotherms showed that the adsorbed amounts of BSA onto PEG-600 surface decreased with GuHCl concentration increment while increased with (NH₄)₂SO₄ concentration increment. FTIR and DSC results showed that the adsorbed BSA molecules obtained more ordered conformation with both GuHCl and (NH₄)₂SO₄ concentration increment. At a given system (a fixed GuHCl and (NH₄)₂SO₄ concentration), with surface coverage increment, although FTIR demonstrated the decrease of ordered secondary structure elements, DSC profiles displayed the enhancement of their thermal stability, which indicates that the total ordered structure obtained still increased.The measured displacement enthalpiesΔH and calculated entropiesΔS and Gibbs energyΔG were all negative. This suggested that the adsorption of denatured BSA onto PEG-600 surface was spontaneous and enthalpy driven process. Based on the analysis of thermodynamic functions, GuHCl produced different effect on the four subprocesses. The affinity of BSA molecules to the surface (subprocesses (a)) and the dehydration between BSA molecules and the surface (subprocesses (c)) were weakened while the conformational gain (subprocesses (b)) and dehydration in BSA molecules (subprocesses (d)) were strengthened. The cooperation of the two opposite effects made the thermodynamic functions -ΔH, -ΔS, -ΔG and their net adsorption fractions -ΔHA, -ΔSA, -ΔGA first increased and then decreased, which exhibited maximum at 0.6 mol·L-1 GuHCl. (NH₄)₂SO₄ promoted the four subprocesses greatly. With (NH₄)₂SO₄ concentration increment, the absolute values of thermodynamic functions and their net adsorption and desorption fractions all increased.