Aggregation of lysozyme and of poly(ethylene glycol)-modified lysozyme after adsorption to silica.

Colloids Surf B Biointerfaces

Department of Chemical Engineering and Department of Biomedical Engineering, Center for Complex Fluids Engineering, Carnegie Mellon University, Pittsburgh, PA 15213, USA.

Published: May 2007

Surface-induced aggregation is a common instability during protein storage, delivery and purification. This aggregation can lead to the formation of fibrils rich in intermolecular beta-sheet structure. Techniques to probe surface-clustering are limited. Here we use protein intrinsic fluorescence and thioflavin T probe fluorescence in a total internal reflection fluorescence (TIRF) sampling geometry to simultaneously monitor the kinetics of adsorption and aggregation for chicken egg lysozyme on a silica surface. We observe a slow surface-induced aggregation process that continues well after the lysozyme adsorption kinetics have plateaued. The rate of surface-induced aggregation is independent of the lysozyme concentration in solution. Consistent with the clustering observed via thioflavin T fluorescence, infrared amide I band spectra also show a 1.5-fold increase in intermolecular beta-sheet content upon lysozyme adsorption. Tryptophan emission spectra show no evidence for any tertiary structural change upon adsorption. Furthermore, we observe that the covalent modification of lysozyme with a single poly(ethylene glycol) (PEG) grafted chain does not inhibit aggregation on the surface, but a second PEG graft significantly inhibits the intermolecular beta-sheet formation.

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Source
http://dx.doi.org/10.1016/j.colsurfb.2007.01.007DOI Listing

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