LAUSR

Arginine, although popularly known as aggregation suppressor additive, has been found to quench protein's structure and function by destabilizing their conformations. Driven by such controversial evidences, in this work we performed a series of atom- istic molecular dynamics simulations of insulin monomer, a biologically active hormone protein, in arginine solution of varying concentrations (0.5 M, 1 M, and 2 M) at am- bient and elevated temperature (400 K) to explore the arginine concentration driven structure-based stability of the protein. Our study reveals that the flexibility of the protein's structure is dependent on the arginine concentration and among all the used solutions, 2 M arginine, a 'neutral crowder' that mimics the cellular environment, can preserve the native folded form of the protein at ambient temperature in an excel- lent manner. Further, while the protein unfolds at 400 K in pure water, this solution worked satisfactory to preserve the protein's folded conformation more firmly, than the other solutions. The replica-exchange MD (REMD) of insulin in 2 M arginine solution further supports the fact. In this aspect an important issue in molecular pharmacology is to identify and recognize the physical origin of the stability of protein, i.e, in this case, how arginine directs the conformational flexibility of the protein and preserves its native folded form. We identified that the exclusion of arginine from the protein sur- face increases the local structuration of water around the protein; thereby preserves its 'biological water' layer and makes the protein more hydrated at 2 M concentration as compared to the other arginine solutions. Additionally, our microscopic investigation on the interactions of protein-solvation layer revealed that the structural heterogene- ity of the protein surface, arising from the differential physicochemical nature of the amino acid residues, controls the favourable formation of sluggish water-arginine mixed solvation layer at higher arginine concentration that helps the protein to maintain its structural rigidity. Importantly, apart from the protein-solvent hydrogen bonding in- teractions, the anion-pi interactions, established between the carboxyl group of arginine and the aromatic amino acid residues of insulin were recognized to facilitate the protein to maintain its native folded form at the experimental temperatures.