Ethanol-Perturbed Amyloidogenic Self-Assembly of Insulin:  Looking for Origins of Amyloid Strains

Abstract

A model cosolvent, ethanol, has profound and diversified effects on the amyloidogenic self-assembly of insulin, yielding spectroscopically and morphologically distinguishable forms of beta-aggregates. The alcohol reduces hydrodynamic radii of insulin molecules, decreases enthalpic costs associated with aggregation-prone intermediate states, and accelerates the aggregation itself. Increasing the concentration of the cosolvent promotes curved, amorphous, and finally donut-shaped forms. According to FT-IR data, inter-beta-strand hydrogen bonding is stronger in fibrils formed in the presence of ethanol. Mechanisms underlying the polymorphism of insulin aggregates were investigated by spectroscopic (CD, FT-IR, and fluorescence anisotropy) and calorimetric (DSC and PPC) methods. The nonmonotonic character of the influence of ethanol on insulin aggregation suggests that both preferential exclusion (predominant at the low concentrations) and direct alcohol-protein interactions are involved. The perturbed hydration of aggregation nuclei appears to be a decisive factor in selection of a dominant mode of beta-strand alignment. It may override unfavorable structural consequences of an alternative strand-to-strand stacking, such as strained hydrogen bonding. A hypothetical mechanism of inducing different amyloid "strains" has been put forward. The cooperative character of fibril assembly creates enormous energy barriers for any interstrain transition, which renders the energy landscape comblike-shaped.

The Same Primary Structure of the Prion Protein Yields Two Distinct Self-propagating States

“…In vitro, the nucleation pathway might depend on the interface between the solution and air or the solution and the wall of the reaction vial (33), whereas interactions of a polypeptide with the surface of biological membranes might regulate the nucleation step in a cell. Using insulin as a model amyloidogenic protein, Winter and co-workers (34) showed that subtle differences in the initial hydration state of a protein at the early stages of fibrillation impose profound consequences for its self-assembling pattern and the physical properties of mature amyloid structures. Our result is consistent with the hypothesis that different nucleation routes lead to well separated energetic minima that correspond to different amyloid strains.…”

Targeting insulin amyloid assembly by small aromatic molecules: Toward rational design of aggregation inhibitors

“…18,19 Moreover, small molecules can also alter the morphology of amyloid fibrils; for instance, ethanol and metalloporphyrins shifted insulin amyloid fibrils into annular structures. 20,21 Thus, small molecules are a powerful tool for understanding the forces effecting the formation and stabilization of amyloidogenic structures.…”

Cytotoxicity of Insulin within its Self-assembly and Amyloidogenic Pathways

“…10,11 Different pathways of non-native aggregation in vivo and in vitro have been described and become manifest in a marked polymorphism of the final aggregate structures, also giving rise to the currently described phenomenon of "strains". [12][13][14][15][16] In living organisms, the native environment of proteins is a complex composition of water, co-solvents, and cosolutes which affect the stability of the native protein fold. 17 Experimental approaches to mimic various cellular environments in vitro and subsequently to characterize the different cytotoxicities of the various protein assemblies often utilize the addition of cosolvents and cosolutes.…”