Insight into the PrP ^C → PrP ^Sc conversion from the structures of antibody-bound ovine prion scrapie-susceptibility variants

Abstract

Prion diseases are associated with the conversion of the ␣-helix rich prion protein (PrP C ) into a ␤-structure-rich insoluble conformer (PrP Sc ) that is thought to be infectious. The mechanism for the PrP C 3 PrP Sc conversion and its relationship with the pathological effects of prion diseases are poorly understood, partly because of our limited knowledge of the structure of PrP Sc . In particular, the way in which mutations in the PRNP gene yield variants that confer different susceptibilities to disease needs to be clarified. We report here the 2.5-Å-resolution crystal structures of three scrapie-susceptibility ovine PrP variants complexed with an antibody that binds to PrP C and to PrP Sc ; they identify two important features of the PrP C 3 PrP Sc conversion. First, the epitope of the antibody mainly consists of the last two turns of ovine PrP second ␣-helix. We show that this is a structural invariant in the PrP C 3 PrP Sc conversion; taken together with biochemical data, this leads to a model of the conformational change in which the two PrP C Cterminal ␣-helices are conserved in PrP Sc , whereas secondary structure changes are located in the N-terminal ␣-helix. Second, comparison of the structures of scrapie-sensitivity variants defines local changes in distant parts of the protein that account for the observed differences of PrP C stability, resistant variants being destabilized compared with sensitive ones. Additive contributions of these sensitivity-modulating mutations to resistance suggest a possible causal relationship between scrapie resistance and lowered stability of the PrP protein.

Molecular Dynamics as an Approach to Study Prion Protein Misfolding and the Effect of Pathogenic Mutations

“…For PrP, the first [8] and most abundant structural information has been obtained by nuclear magnetic resonance techniques (NMR), which has resulted in experimentally derived models of PrP of a wide variety of species [64][65][66][67][68][69][70]. For human and sheep PrP, structures determined by X-ray crystallography have also been reported, both with and without antibodies bound [71][72][73][74][75]. The overall structure of PrP that has emerged from these studies is largely identical for the different species and conditions.…”

Probing the origins, diagnosis and treatment of amyloid diseases using antibodies

“…One of these ways is to generate antibodies against diverse epitopes of the native protein (antibodies 1-3 in Fig. 1) and then to compare their binding properties before and after the conformational changes that result in the formation of amyloidogenic intermediates [47]. The region(s) of the protein where such changes occur, and those where the native structure is conserved, can then be mapped, at least in outline.…”

“…This latter strategy has been used to generate both polyclonal and monoclonal antibodies specific to the pathogenic scrapie form of the prion protein (PrP Sc ) [52,53]. Using a combination of these approaches, a range of recombinant antibodies has been generated and used to map the conformational changes associated with the formation of PrP Sc [47,[52][53][54][55][56][57] or to probe the conformation of the normal cellular form of the prion protein (PrP C ) on the cell-surface [58].…”

Misfolding of the prion protein: linking biophysical and biological approaches

“…More recently, high resolution crystallographic structures were established for sheep PrP variants [32,43]. They showed the same global fold as other mammalian species determined by NMR.…”

“…They showed the same global fold as other mammalian species determined by NMR. However, these high resolution structures led to establish the molecular bases of the difference in the previously described energetic unfolding [32,98]. Furthermore, the use of an antibody in the crystallization trials helps to identify the segment of ovine PrP (OvPrP) with an unchanged secondary structure in the PrP C -toPrP Sc conversion.…”