THE EFFECT OF SOLUTION PH ON THE STRUCTURE OF LYSOZYME LAYERS ADSORBED AT THE SILICA-WATER INTERFACE STUDIED BY NEUTRON REFLECTION

We have studied the structure of lysozyme layers adsorbed at the silic a-water interface using specular neutron reflection. The effect of pH on the adsorbed lysozyme layer was examined by manipulating the pH in two different cycles at two constant lysozyme concentrations of 0.03 a nd 1.0 g dm(-3); the first cycle was started at pH = 4 followed by pH = 7 and then 8, before returning to 4; the second cycle was started at pH = 7 followed by a decrease to 4 and then back to 7. The neutron re flectivity profiles showed no hysteresis in either adsorbed amount or structure. There was less adsorption at pH = 4 than at pH 7 for both l ysozyme bulk concentrations. No variation of the reflectivity with tim e was found at the experimental resolution of about 5 min per measurem ent. The lysozyme structure at the interface at pH = 4 and pH = 7 was determined from reflectivity profiles at different isotopic compositio ns of the water. The thickness of the adsorbed layer at the lower conc entration of 0.03 g dm(-3) was found to be 30 +/- 2 Angstrom, suggesti ng sideways-on adsorption of the ellipsoidally shaped protein. At the higher concentration of 1.0 g dm(-3) the thickness of the layer was fo und to be 60 +/- 2 Angstrom, suggesting bilayer adsorption with side-o n orientation in each layer. These observations disagree with literatu re results from surface force and ellipsometric measurements which sug gest that a side-on monolayer of 30 Angstrom thickness is formed at di lute bulk concentrations, which switches to end-on adsorption of 45 An gstrom thickness as the bulk concentration increases, eventually reach ing a bilayer of 90 Angstrom thickness when the bulk lysozyme concentr ation is further increased. The neutron measurements indicate that the adsorbed amount and the orientation of the globular protein are deter mined by the electrostatic repulsion between the lysozyme molecules wi thin the layer.