Assemblages of prion fragments: novel model systems for understanding amyloid toxicity

Abstract

We report the conformational and toxic properties of two novel fibril-forming prion amyloid sequences, GAVVGGLG (PrP(119–126)) and VVGGLGG (PrP(121–127)). The conformational preferences of these fragments were studied in differing microenvironments of TFE/water mixtures and SDS solution. Interestingly, with an increase in TFE concentration, PrP(119–126) showed a helical conformational propensity, whereas PrP(121–127) adopted a more random coil structure. In 5% SDS, PrP(119–126) showed more α-helical content than in TFE solution, and PrP(121–127) exhibited a predominantly random coil conformation. However, both peptides took a random coil conformation in water, and over time the random coil transformed into a β-sheet structure with a significant percentage of helical conformation and β-turn structure in PrP(119–126) and PrP(121–127), respectively, as observed with CD spectroscopy. The aged fibrils of PrP(119–126) were insoluble in SDS, and PrP(121–127) was extractable with SDS solution. These fibrils were characterized by transmission electron microscopy. Both PrP(119–126) and PrP(121–127) formed stable monolayer's consisting of multimeric assemblages at the air–water interface. Monomeric PrP(119–126) was more toxic to astrocytes than the control Aβ peptide; however, the fibrillar form of PrP(119–126) was less toxic to astrocytes. PrP(121–127) elicited moderate toxicity in both soluble and fibrillar forms on astrocytes. Furthermore, quenching experiments using acroyl-labeled PrP(119–126) and PrP(121–127) with eosin-labeled synaptosomal membrane revealed that these prion fragments bind to anion-exchange protein. The binding of PrP(119–126) and PrP(121–127) with a membrane microdomain (lipid raft) was also analyzed using pyrenated derivatives. We conclude that the formation of PrP(119–126) and PrP(121–127) fibrils is a concentration-dependent process that involves coil to sheet conversion with aging. PrP(119–126), the sequence with intrinsic helical propensity, is more toxic in monomer form, and the fibril formation in this case seems to be protective to cells. For PrP(121–127), the SDS-soluble fibrils are more cytotoxic, indicating that a higher order assemblage structure is required for cytotoxic activity of this peptide.