(269h) Kinetics of Unfolding of Adsorbed Beta Lactoglobulin on Silica Nanoparticle Surface
Fluorescence spectra of aromatic residues of adsorbed beta lactoglobulin molecules on surface of silica nanoparticle of 90 nm diameter when excited at 280 nm was measured as a function of time. Unfolding of the protein molecule resulted in an increase in the intensity of the spectra with time because of exposure of more aromatic residues. The intensity of fluorescence spectra was measured for native beta lactoglobulin and protein with different extents of denaturation(with guanidium chloride) which were used as reference in the conversion of fluorescence intensity to extent of unfolding. The extent of unfolding was more at lower surface concentrations, lower ionic strengths and near pI. The rate as well as the extents of unfolding were found to be smaller at higher surface concentrations possibly due to steric hinderence of neighboring adsorbed molecules on the surface. Since pH 5 is closer to pI of beta lactoglobulin, the adsorbed protein molecule experienced smaller electrostatic interactions with neighboring molecules thus allowing more unfolding. At pH 7, however, the adsorbed molecules experienced stronger electrostatic repulsion which hindered unfolding. The difference between unfolding at two pH values was found to be more pronounced at lower surface concentrations. At higher ionic strength, the electrostatic repulsion between neighboring adsorbed molecules was suppressed because of shielding of charges thus enabling more unfolding. Interestingly, the difference in extent of unfolding was significant even at lower surface concentrations. A phenomenological model for unfolding was proposed which accounted for adsorption of protein from the bulk followed by unfolding at the surface. The adsorbed protein molecule was visualized as being in native, intermediate, unfolded and dimer states. The model was shown to describe qualitatively the experimental behavior.