On the mechanism of alpha-helix to beta-sheet transition in the recombinant prion protein

It is believed that the critical event in the pathogenesis of transmissible spongiform encephalopathies is the conversion of the prion protein from an alpha -helical form, PrPC, to a beta -sheet-rich conformer, PrPSc. Recentl y, we have shown that incubation of the recombinant prion protein under mil dly acidic conditions (pH 5 or below) in the presence of low concentrations of guanidine hydrochloride results in a transition to PrPSc-like beta -she et-rich oligomers that show fibrillar morphology and an increased resistanc e to proteinase K digestion [Swietnicki, W., Morillas, M, Chen, S., Gambett i, P., and Surewicz, W. K. (2000) Biochemistry 39, 424-431]. To gain insigh t into the mechanism of this transition, in the present study we have chara cterized the biophysical properties of the recombinant human prion protein (huPrP) at acidic pH in the presence of urea and salt. Urea alone induces u nfolding of the protein but does not result in protein self-association or a conversion to beta -sheet structure. However, a time-dependent transition to beta -sheet structure occurs upon addition of both urea and NaCl to huP rP, even at a sodium chloride concentration as low as 50 mM. This transitio n occurs concomitantly with oligomerization of the protein. At a given prot ein and sodium chloride concentration, the rate of monomeric alpha -helix t o oligomeric beta -sheet transition is strongly dependent on the concentrat ion of urea. Low and medium concentrations of the denaturant accelerate the reaction, whereas strongly unfolding conditions are not conducive to the c onversion of huPrP into an oligomeric beta -sheet-rich structure. The prese nt data strongly suggest that partially unfolded intermediates may be invol ved in the transition of the monomeric recombinant prion protein into the o ligomeric scrapie-like form.