Prevention of insulin fibrillation by biocompatible choline-amino acid based ionic liquids: Biophysical insights

Abstract

We have synthesized biocompatible ionic liquids (ILs) with choline as cation and amino acids as anions to explore their potential towards prevention of fibrillation in insulin and the obtain corresponding mechanistic insights. This has been achieved by examining the effect of these ILs on insulin at the nucleation, elongation and maturation stages of the fibrillation process. A combination of high sensitivity isothermal titration calorimetry (ITC) and differential scanning calorimetry (DSC) have been employed along with spectroscopy and microscopy to evaluate interaction of the ILs at each stage of fibrillation quantitatively. Choline glycinate is observed to provide maximum stabilization to insulin compared to that provided by choline prolinate, choline leucinate, and choline valinate. This increased thermal stabilization has direct correlation with the extent of reduction in the fibrillation of insulin by ILs determined using Thioflavin T and 8-anilinonaphthalene sulfonate based fluorescence assays. ITC has permitted understanding nature of interaction of the ILs with the protein at different fibrillation stages in terms of standard molar enthalpy of interaction whereas DSC has enabled understanding the extent of reduction in thermal stability of the protein at these stages. These ILs are able to completely inhibit formation of insulin aggregates at a concentration of 50 mM. Stabilization of proteins by ILs could be explained based on involvement of preferential hydration process. The work provides biocompatible IL based approach in achieving stability and prevention of fibrillation in insulin.