Structural organization of brain-derived mammalian prions examined by hydrogen-deuterium exchange

Abstract

One of the mysteries in prion research is the structure of the infectious form of mammalian prion protein PrP(Sc). Here we used mass spectrometry analysis of hydrogen-deuterium exchange to examine brain-derived PrP(Sc). Our data indicate that, contrary to popular models, prion-protein conversion involves refolding of the entire region from residue ~80-90 to the C-terminus, which in PrP(Sc) consists of β-strands and relatively short turns and/or loops, with no native α-helices present.

Figure 1

Deuterium incorporation for peptic fragments derived from different types of misfolded prion protein aggregates. (a) PrP^Sc from wild-type mice infected with 22L strain of scrapie after 240 hours of exchange. (b) PrP^Sc from transgenic GPI⁻ mice infected with 22L strain of scrapie after 5 min and 240 hours of exchange. (c) PrP^Sc from transgenic GPI⁻ mice infected with Chandler strain of scrapie after 240 hours of exchange. (d) PrP^Sc from transgenic GPI⁻ mice infected with ME7 strain of scrapie after 240 hours of exchange. (e) Amyloid fibrils formed from the recombinant mouse prion protein 89–231 after 240 h of exchange. Error bars indicate s.d.

Figure 2

Schematic representation of prion protein secondary structure in the ß-helix (B) and spiral (S) models of PrP^Sc. Black arrows, curved lines and solid lines represent ß-strands,α-helices and loops/unordered segments, respectively. Colour bars immediately below the sequence represent the percentage of deuterium incorporation for GPI⁻22L PrP^Sc after 240 hours of exchange.

Figure 3

Pairwise comparison of difference in deuterium labeling after 240 h exchange for different PrP^Sc strains. (a) 22L and Chandler GPI PrP^Sc. (b) 22L and ME7 GPI⁻PrP^Sc. (c) Chandler and ME7 GPI⁻PrP^Sc. Data are based on 3–5 experiments using two different preparations of 22L and Chandler GPI⁻PrP^Sc and a single preparation of ME7 GPI⁻PrP^Sc. Error bars indicate s.d.