Characterization of secondary and tertiary conformational changes of beta-lactoglobulin adsorbed on silica nanoparticle surfaces.
Nanoparticles possess unique properties as a result of their large surface area per unit volume and therefore can be functionalized by the immobilization of enzymes for a variety of biosensing applications. Changes in the tertiary conformation of beta-lactoglobulin adsorbed on 90 nm silica nanoparticles with time were inferred using tryptophan fluorescence and Fourier transform infrared spectroscopy (FTIR) for different surface concentrations, temperature, pH, ionic strength, and 2,2,2-trifluoroethanol (TFE) and dithiothreitol (DTT) concentrations. Rapid initial unfolding followed by a much slower rate at longer times was observed, with the extent of unfolding being higher at lower surface concentrations, higher ionic strengths, higher temperature, higher TFE and DTT concentrations, and pI. The effect of temperature on the unfolding of adsorbed protein on the nanoparticle surface was similar to that in the bulk even though the extent of unfolding was higher for adsorbed protein molecules. The results of the extent of change in tertiary conformation using FTIR as indicated by the change in the ratio of amide II'/amide I were consistent with those obtained by tryptophan fluorescence whereas the rates of conformational changes given by FTIR were found to be much faster. Circular dichroism (CD) spectra showed that altering the surface concentration by itself did not change the secondary structure of beta-lactoglobulin on the surface. TFE was found to increase the alpha helix content at the expense of the fraction of the beta sheet, whereas the beta sheet was converted to an unordered conformation in the presence of DTT.
Wu
Xiaoyu
X
Biochemical and Food Process Engineering, Department of Agricultural and Biological Engineering, Purdue University, West Lafayette, Indiana 47907, USA.
Narsimhan
Ganesan
G
D000327:Adsorption; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007782:Lactoglobulins / Q000737:chemistry* / Q000378:metabolism; D053758:Nanoparticles / Q000737:chemistry*; D009994:Osmolar Concentration; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012822:Silicon Dioxide / Q000737:chemistry*; D017550:Spectroscopy, Fourier Transform Infrared; D013499:Surface Properties; D013696:Temperature
D007782:Lactoglobulins; D012822:Silicon Dioxide