Investigating Reliable Conditions for HEWL as an Amyloid Model in Computational Studies and Drug Interactions

A number of conformational diseases in humans have been associated with protein/peptide fibrillation known as amyloid. Although extensive studies have been conducted in understanding the molecular basis of amyloid formation, a detailed mechanism is still missing. Experimentally, HEWL (hen egg white lysozyme) has been exploited ubiquitously as a model protein for amyloid fibrillation and drug inhibition. However, computational studies investigating fibril formation of HEWL have been a difficult task to perform mainly due to high stability of lysozymes and the absence of crystal structures of HEWL fibril oligomers. In this study, we have examined various conditions of HEWL amyloid formation computationally; the results indicated that, at high concentration of ethanol (90%), significant unfolding of the protein was apparent. Higher values for RMSD, solvent accessibility, and solvent diffusion into the core, as well as conversion of native α-helical structures to random coils, were detected in the ethanol solution. REMD (replica exchange molecular dynamics) analysis demonstrated that the presence of ethanol significantly altered the minimum structure of HEWL into partially unfolded states. It has been observed that unfolding of the protein was initiated from the C-terminal region, exposing the protein to the solvent. The interaction of previously known anti amyloid drug (RS-0406) with HEWL was analyzed in high concentration of ethanol both in silico and in vitro. The results demonstrated that the drug was able to attenuate HEWL unfolding and fibrillation both experimentally and computationally. Computational studies provided detailed interactions explaining the inhibitory effect of the drug in this model. Most importantly, a mechanism of drug inhibition was purported based on a bridge formed by the drug that stabilized the C-terminus. All in all, a computational model of HEWL amyloid formation was attained which can be employed to assess inhibitory effects of antiamyloid drugs in a reasonable processing time.