Protein structural perturbation and aggregation on homogeneous surfaces

Abstract

We have demonstrated that globular proteins, such as hen egg lysozyme in phosphate buffered saline at room temperature, lose native structural stability and activity when adsorbed onto well-defined homogeneous solid surfaces. This structural loss is evident by α-helix to turns/random during the first 30 min and followed by a slow α-helix to β-sheet transition. Increase in intramolecular and intermolecular β-sheet content suggests conformational rearrangement and aggregation between different protein molecules, respectively. Amide I band attenuated total reflection/Fourier transformed infrared (ATR/FTIR) spectroscopy was used to quantify the secondary structure content of lysozyme adsorbed on six different self-assembled alkanethiol monolayer surfaces with -CH3, -OPh, -CF3, -CN, -OCH3, and -OH exposed functional end groups. Activity measurements of adsorbed lysozyme were in good agreement with the structural perturbations. Both surface chemistry (type of functional groups, wettability) and adsorbate concentration (i.e., lateral interactions) are responsible for the observed structural changes during adsorption. A kinetic model is proposed to describe secondary structural changes that occur in two dynamic phases. The results presented in this article demonstrate the utility of the ATR/FTIR spectroscopic technique for in situ characterization of protein secondary structures during adsorption on flat surfaces. © 2005 by the Biophysical Society.