Investigation of conformational changes of protein adsorbed on silica nanoparticle surface

Xiaoyu Wu, Purdue University

Abstract

Nanoparticles possess unique properties due to large surface area per unit volume and therefore can be functionalized for a variety of biosensing applications. It is important to characterize the unfolding of proteins/enzymes at surfaces because immobilization of proteins/enzymes can lead to partial or complete loss of its activity. Changes in tertiary conformation of protein adsorbed on silica nanoparticles was measured using tryptophan fluorescence and Fourier Transform Infrared Spectroscopy and changes in secondary conformation was measured by Circular Dichroism (CD) spectrum. A rapid initial unfolding of β lactoglobulin adsorbed on the silica nanoparticles surfaces followed by a much slower rate at larger times was observed with the extent of unfolding being higher at lower surface concentrations, higher ionic strengths, higher 2,2,2,-trifluoroethanol (TFE) and Dithiothreitol (DTT) concentrations and at pI. For lysozyme adsorbed on the silica nanoparticle surface, a rapid initial unfolding, followed by a slower refolding and subsequent unfolding, were observed. CD spectra revealed that surface concentration and TFE have the effect on the secondary structure of lysozyme on the surface, whereas such an effect is not found on β lactoglobulin. The difference between the lowest energy structures of TC5b and TC5c obtained by all atom molecular dynamics (MD) simulation lies in the absence of 310 helix for the latter because of the absence of electrostatic interaction. Cecropin P1 C is found to be more stable than Cecropin P1 in solution as shown by the analysis of α helix and native contacts at different temperatures. A coarse grain MD simulation algorithm was developed in which the polypeptide backbone and sidechains were replaced by beads of equivalent interactions for the prediction of tertiary conformation. Coarse grain (CG) algorithm revealed that lysozyme molecule adsorbed on silica surface is more unfolded with a less compact tertiary structure compared to that in the solution. Higher ionic strength resulted in a more extended structure for lysozyme on silica surface. All atomic simulation results showed a loss of the tertiary structure of protein (Trp-cage and Cecropin P1 C) due to the interaction with silica surface, while a certain secondary structure is found to be still retained.

Degree

Ph.D.

Advisors

Narsimhan, Purdue University.

Subject Area

Food Science|Bioinformatics|Biophysics

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