THE DENATURATION OF LYSOZYME LAYERS ADSORBED AT THE HYDROPHOBIC SOLIDLQUID SURFACE STUDIED BY NEUTRON REFLECTION/

We have studied the adsorption of lysozyme layers at a hydrophobic sil icon water interface using specular neutron reflection. The hydrophobi c surface was obtained by self-assembly of a densely packed monolayer of octadecyltrichlorosilane (OTS) onto the natural silica layer on the smooth surface of a (111) silicon block. The effect of pH on the adso rbed lysozyme layer was examined at a constant lysozyme concentration of 0.03 g dm(-3) and at a constant ionic strength of 0.02 M. Reflectiv ity profiles at different pH show that adsorption is irreversible with respect to pH, the composition and structure of the final layer being dependent on the route by which the pH was achieved. The adsorbed pro tein layer was found to divide into approximately two regions, a dense ly packed thin layer next to the OTS surface and a diffuse thicker lay er extending into the bulk solution. None of the dimensions of this st ructure corresponds to those of the globular protein in solution, sugg esting that, unlike its adsorption at the hydrophilic silica/water int erface, lysozyme is denatured at the OTS/ water surface. The irreversi ble adsorption is explained by the combined interaction of the hydroph obic attraction of the hydrophobic fragments in lysozyme to the OTS su rface and electrostatic repulsion within the adsorbed layer. The hydro phobic surface induces the exposure of hydrophobic fragments from the lysozyme assembly. The thickness of the dense layer suggests that the denatured protein adsorbs in the form of peptide chains with the hydro phobic amino acid side chains attached to the OTS surface with the hyd rophilic side chains extending into the bulk solution. Since lysozyme is more stable at pH 7 than at pH 4, the difference in initial adsorpt ion is dominated by the greater relative stability of lysozyme to dena turation at the higher pH. A change of pH from 7 to 4 reduces the stab ility of the protein to unfolding and results in more adsorption than when the pH is changed in the opposite direction. Solution pH also aff ects the net charges within the hydrophilic tail region and the struct ural distribution of the tail region was found to vary with pH, (C) 19 98 Academic Press.