Ellipsometry and TIRF studies of enzymatic degradation of interfacial proteinaceous layers

Ellipsometry and total internal reflectance fluorescence spectroscopy (TIRF) have been employed to investigate the layer structure of gelatin adsorbed from aqueous solutions onto silica/glass and methylated silica/glass, as well as the effects of addition of the proteolytic enzymes krillase and trypsin, in relation to temperature, enzyme concentration, and enzymatic activity. The results for the hydrophilic substrates show that homogeneous and heterogeneous exchange occurs readily, as does autolysis of trypsin at the interface. At the hydrophobic substrates, the effect of exchange is limited and a residual gelatin fraction is present at the interface throughout. The interfacial behavior of gelatin above and below the helix formation temperature (Thelix) shows that more extended surface layers are formed at both substrates below Thelix. At the hydrophilic substrates, the higher adsorbed layer thickness below Thelix is mainly due to the adsorption of more gelatin than at the higher temperature, whereas, at ...