Revisiting the conundrum of trehalose stabilization

Abstract

Protein aggregation and loss of protein's biological functionality are manifestations of protein instability. Cosolvents, in particular trehalose, are widely accepted antidotes against such destabilization. Although numerous theories have been promulgated in the literature with regard to its mechanism of stabilization, the present scenario is still elusive in view of the discrepancies existing in them. To this end, we have revisited the conundrum and attempted to rationalize the mechanism by conducting thorough investigation of the effect of trehalose on the native, partially unfolded and denatured states of protein "Lysozyme" by means of molecular dynamic (MD) simulations under different temperature and concentration regimes. Two-dimensional contour plots along with principal component analysis suggest that trehalose molecules offer on-pathway stabilization unaltering the principal direction of protein's motion, although it slows down protein dynamics so that the protein gets trapped in the homogeneous ensemble of conformations closer to the native state. Free energy landscape reveals higher population of native compared to intermediate and denatured states. Delphi results and calculation of the preferential interaction parameter demonstrate that this relative stabilization of the native state can be ascribed to be the consequence of favourable interactions of trehalose with side chains of certain loci on the protein surface encompassing polar flexible residues. Stability of protein results from the observed difference in binding affinity of trehalose for native and denatured states of protein. Our findings are at variance with the common conception of relative destabilization of the denatured state. Rather, we provide evidence for relative stabilization of the native state. This stabilization is due to interplay of protein-trehalose, water-trehalose, water-water, protein-water and trehalose-trehalose interactions.

Collaborative routes to clarifying the murky waters of aqueous supramolecular chemistry

“…Beyond the nonspecific binding of glycerol, does trehalose stabilization of proteins and its role in anhydrobiosis (a dormant state induced by drought whereby an organism becomes almost completely dehydrated) involve any degree of specificity? 123 Can small polyol additives be used to mimic bound waters at specific sites on a protein surface? And if so, which residues are key for selective recognition.…”

Practical insights on enzyme stabilization

“…Enzymes are protein molecules consisting of folded polypeptide chains of amino acids that are essential to perform an array of biological functions. The order of these amino acids in a protein determines its tertiary structure through molecular geometry and intramolecular chemical interactions [7,8]. Depending on the amino acid composition, these proteins can incorporate both acidic and basic functional groups, which play an important role in their structure.…”

More than a Confinement: “Soft” and “Hard” Enzyme Entrapment Modulates Biological Catalyst Function

“…The larger flexibility in saccharide confinement makes porosity not the primary measure of the confining ability: polysaccharide matrices with defined porosity do exist, but the macromolecular scaffold is in any case less rigid and allows a certain degree of adaptability, allowing also the hosting of macromolecules with dimensions or shapes not properly matching those of the pores. In some cases, specific interactions can be observed between the sugar hydroxyls and definite parts of the embedded protein, resulting in an increased efficiency in preservation [15,16]. Low MW saccharide matrices (oligo or disaccharides) lack entirely the concept of a pore, as they are true amorphous matrices without a regular supramolecular organization.…”

“…Reducing the water content, a few direct protein-trehalose interactions start to emerge, indicating that the water replacement and water entrapment hypotheses are not mutually exclusive, but their occurrence depends only on external parameters, as hydration or sugar/protein ratio [163]. MD simulations showed also that trehalose could form patches around the protein, reducing the protein backbone flexibility [16,164].…”