X-ray Diffraction of Scrapie Prion Rods and PrP Peptides

Abstract

Certain neurodegenerative diseases in humans and animals are caused by small proteinaceous infectious particles called prions. Limited proteolysis and detergent extraction of the prions containing PrP Scgenerate prion rods that are composed of a polypeptide having an apparent molecular mass of 27 to 30 kDa. This polypeptide, termed prion protein PrP 27–30, has a ragged N terminus that begins at about residue 90, but retains scrapie infectivity. Moreover, the findings in a patient having an inherited prion disease of a truncated PrP with its C terminus at residue 145 suggest that the residues 90 to 145 may be of particular importance in the pathogenesis of prion diseases. To determine the three-dimensional organization of prion rods and to identify the core region involved in amyloid formation, we recorded X-ray diffraction patterns from rods purified from scrapie-infected Syrian hamster (SHa) brains which contain PrP 27–30, and from synthetic SHaPrP peptides. Three peptides were studied corresponding to residues 113 to 120 (peptide A8A, an octamer composed of glycines and alanines), 109 to 122 (H1, the first predicted α-helical region of PrP C), and 90 to 145 (a 56 residue peptide containing both H1 and the second predicted α-helical region of PrP C, H2). Electron microscopy, carried out in parallel with the X-ray measurements, revealed that all the samples formed linear polymers which were ∼60 to ∼200 Å wide, with fibrillar or ribbon-like morphology. Gels and dried preparations of prion rods gave X-ray patterns that indicated a β-sheet conformation, in which the hydrogen bond distance was 4.72 Å and the intersheet distance was 8.82 Å. For the three PrP peptides, the intersheet spacings varied widely, owing to the side-chains of the residues involved in the formation of the β-sheet interactions, i.e., 5.13 Å for A8A, 5.91 Å for lyophilized H1, 7.99 Å from solubilized and dried H1 and 9.15 Å for the peptide SHa 90–145. The intersheet distance of PrP 27–30 was thus within the observed range for the peptides, and suggests that the amyloidogenic core of PrP is closely modeled by the peptide SHa 90–145.