Solution thermodynamics and interaction studies of protein-osmolyte mixtures by NMR

This PhD thesis focuses on the role of osmolytes in stabilizing/destabilizing proteins involved in amyloidoses, i.e. diseases developing as a consequence of protein misfolding. We have chosen to study three amyloidogenic systems viz.; the eye-lens protein α-crystallin which upon aggregation and precipitation give rise to cataract, a fragment from the Amyloidβ-peptide (Aβ) a major constituent of the Alzheimer's plaque and β2-microglobulin (β2m) the protein responsible for dialysis related amyloidosis. Using NMR experiments we have tried to characterize the type of interactions of the first two systems with a well-studied protein-stabilizing osmolyte, carnosine. Next, we have looked at the unfolding free energy landscape of β2m in the presence of the procteting osmolyte carnosine and the denaturing osmolyte heparan sulfate by using hydrogen-deuterium NMR exchange experiments. Our results from interaction studies suggests the presence of transient protein:osmolyte interactions which in the case of α-crystallin was very efficient and could be traced up to very high concentrations of the osmolyte (a protein-osmolyte ratio 1:1000). The results from isotope exchenge experiments throws light on the global and local unfoldin events occurring in the model protein β2m in the presence and absence of two osmolytes