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Reversible unfolding of infectious prion assemblies reveals the existence of an oligomeric elementary brick

Fig 1

Sequential unfolding and refolding of PrPSc trapped an elementary conformer.

(A) Overview of the procedure for unfolding and refolding native PrPSc. (B) The effect of an increasing urea concentration on the quaternary structure of PrPSc was analysed based on SV. The superposition of the SV profile of PrP after urea treatment (the blue-to-red scale corresponds to 0 to 6 M urea) emphasizes the quaternary structure transition with an isobestic point (arrow) from highly polydisperse assemblies to a smaller and monodispersed conformer. The black line indicates the SV profile of PrPC in 6 M urea (PrPU). (C) Similar experiments as in (B), with a refolding step via dialysis to remove urea (the blue-to-red scale corresponds to 0 to 6 M urea treatment before dialysis). The SV profile of PrP reveals the formation of two sets of assemblies termed rf1PrP and rf2PrP. The SV profile of PrPC treated with urea and refolded via dialysis is represented as a black line with an intensity adjustment. (D) The evolution of the sedimentogram centroid of PrPSc unfolding (B) and refolding (C) as function of the urea concentration revealed a sigmoidal shape for the unfolding and refolding step, suggesting a cooperative phenomenon. (E) Size exclusion chromatography profile of partially purified suPrP263K (in red), purified [31] suPrP263K (in green) and purified PrPC unfolded in 6 M urea (in black). As shown, partially purified and purified suPrP263K presented an oligomeric structuration (for details, see Materials and Methods and appendix S2 and S3 Appendices). (F) scheme summarising the PrPSc unfolding process induced by urea and the formation of suPrP. This last refolds into rfPrP after urea removal by dialysis.

Fig 1