Spectroscopic Studies On Pressure-induced Conformational Changes Of Biomolecules In Aqueous Solution

Protein aggregation playssignificant but, to date, undefined roles in severalhuman diseases. The identification of protein-folding intermediates has becomeincreasingly important with the finding that they could act as precursors to themisfolded and aggregated species that are related to several neurodegenerativediseases, such as Alzheimer’s, Parkinson’s, transmissible encephalopathies, sometypes of cancer and systemic amyloidogenic diseases. Hydrostatic pressure is a usefulway to perturb the tertiary and quaternary structures of proteins based on volumechanges associated with the presence of distinct solvent-accessible regions andcavities in different conformational states.Pressure as well as temperature induced effects in proteins are closely connectedwith the presence of water. Changes in the interaction with water (hydration) and theimperfect packing effects in the interior (void volume and cavities) contribute greatlyto the structural and dynamic variations of proteins in aqueous solutions.Pressure is able to change the shape of a protein, extremely high pressure can leadto unfolding of the protein, but lower pressures cause only elastic distortion of thesecondary and tertiary structures. In order to clearly explain the relationship betweenstructure changes of water and protein, we chose four typical proteins, lysozyme,human serum albumin (HSA), ubiquitin and RNase A, these have different secondarystructures and space conformations. We investigated pressure-induced structurechanges of four proteins in aqueous solution from0.1to740MPa by using infrared spectroscopy. Both the hydration and minor structural changes of proteins have beenrevealed clearly by principal component analysis (PCA) and moving-windowtwo-dimensional. With regard to the pressure induced structure changes of lysozyme,HSA, ubiquitin, and RNase A, it became obvious that the characterized amide I bandof protein shifted to lower wavenumbers with a common transition at200MPa,indicating that pressure induced a hydration increase at the lower pressure region.This is similar to the pressure induced structure change of pure water, and it ispossible that the structure change in the proteins studied is caused by the structurechange of water. Moreover, the change processes for the proteins were found not to betwo-state processes, as several fluctuations along the pressure change were observedaccording to the analytical results of the PCA and MW2D analyses.These studies on pressure induced conformational changes in proteins showed thateach protein experienced an effect on the unfolding that wa sdriven by its individualamino acid sequence. Choosing either the isozymes of an enzyme from one species orthe same proteins from several different species as studying system is a goodpossibility to exclude the effect of conformation, as they are high homology and albeitmay response to pressure very identically. In addition, to investigate the effect of theconformation but the amino acid sequences on unfolding, it is necessary to find anideal system where the sequence is the same but the conformation is different. In thepresent study, we used FT-IR spectra and two-dimensional correlation analysis toinvestigate systematically the pressure-induced structural changes in three initialconformations of Poly（L-lysine） in aqueous solution; Furthermore, we examined theeffects of different chain lengths. Three important conclusions can be drawn from thepresent investigation. First, high pressure is able to induce conformational changes ina70kDa Poly （L-lysine）. With increasing pressure, significant changes in theα-helix, β-sheet, and random coil are observed at around300,250, and850MPa,respectively. The pressure require to produce these transitions is most likely related tothe local isoelectric point of polylysine. Moreover, of the three conformers, theβ-sheet appears to be the most sensitive and the random coil is the least sensitive to pressure. A second conclusion, which should be emphasized, is that all the initialconformers underwent pressure-ind uced conformational changes to assume the samefinal structure, the α′-helix; this structure may be a new, reorganized, highly hydratedα-helical structure that is adapted to high pressures. Our third conclusion is that theproduct of the pressure-induced β-sheet toα-helix transition differs from the initialα-helix.Pressure-induced fluorescence enhancements of AuNCs@BSA are investigated inthe present study. On the basis of the combined experimental information presentedin this paper, we can conclude that it is the protected ligand other than the corerearrangement play major role for the fluorescence enhancement of AuNCs@BSA.BSA adopts a more flexible conformational state at the boundary surface of AuNCs asa result of different conformational changes way in the bioconjugates from nativeBSA. The pressure-induced disruption of salt bridges, the enhanced hydration ofnewly exposed nonpolar and polar residues, and especially the disruption ofhydrophobic interactions of ligand influence the optical property of gold nanoclustersgreatly. Albeit these results are in agreement with the findings reported previouslyfor the small molecule-protected luminescent gold nanoclusters, the present study stilllays the foundation for improving the NCs luminescence by adjusting theconformation of protein ligand. In addition, the reported environment-sensitive opticalproperties of metal NCs have enabled the development of new techniques forsensitive chemical and biological sensing.