The double-edged influence of guanidinium chloride on protein aggregation

BALLATORE, M; DELFINO J.M.; CURTO L.M.

Congreso; 20th IUPAB Congress, 45th Annual SBBf Meeting, and 50th Annual SBBq Meeting; 2021

Due to its ability to form micelle-like clusters, trifluoroethanol (TFE) promotes conformational changes that can trigger amyloid formation. Unlike classical denaturants, TFE favors protein aggregation mainly at moderate concentrations (~30% v/v). Accordingly, our protein model IFABP (intestinal fatty acid binding protein) shows amyloid-like aggregation above 15% v/v. As a straightforward correlation has been generally postulated between stability and amyloid propensity, our aim was to address the influence of sub-aggregating TFE concentrations on IFABP stability. Urea or guanidinium hydrochloride (GdmCl)-induced unfolding transitions were monitored by fluorescence and circular dichroism (CD) spectroscopy. Insights on the process arise from the comparison of the shape and intensity of the full set of CD spectra obtained from thermal ramps performed with a Chirascan V-100 (Applied Photophysics). With urea, IFABP can be assimilated to a 2-stage system and its stability remains unchanged. At variance, with GdmCl, the appearance of amyloid-like aggregation becomes more evident as TFE concentration increases. Temperature-induced denaturation profiles show that both additives diminish stability. Whereas a concomitant increase of amorphous aggregation occurs upon heating in the presence of TFE, no aggregation takes place with GdmCl. Conversely, when both additives are present, amyloid-like aggregation initiates immediately. In summary, two aggregation pathways might happen: amorphous or amyloid-like. Upon heating, low TFE concentrations promote the first. If GdmCl is also present the second route takes over. The explanation must reconcile the effects of additives on both the protein and solvent structures. Briefly, TFE desolvates the protein, a process further reinforced by heat. Although GdmCl might prevent thermal aggregation by solubilizing non-native states, this same phenomenon could favor amyloid aggregation. On top of that, the electrolytic-induced segregation of TFE might contribute to the development/stabilization of TFE clusters that might reach a large enough size to act as nucleation-inducing interfaces, thus leading to the observed amyloid aggregation.