Abstract
The structure of the infectious form of prion protein, PrP(Sc), remains unclear. Most pure recombinant prion protein (PrP) amyloids generated in vitro are not infectious and lack the extent of the protease-resistant core and solvent exclusion of infectious PrP(Sc), especially within residues ∼90-160. Polyanionic cofactors can enhance infectivity and PrP(Sc)-like characteristics of such fibrils, but the mechanism of this enhancement is unknown. In considering structural models of PrP(Sc) multimers, we identified an obstacle to tight packing that might be overcome with polyanionic cofactors, namely, electrostatic repulsion between four closely spaced cationic lysines within a central lysine cluster of residues 101-110. For example, in our parallel in-register intermolecular β-sheet model of PrP(Sc), not only would these lysines be clustered within the 101-110 region of the primary sequence, but they would have intermolecular spacings of only ∼4.8 Å between stacked β-strands. We have now performed molecular dynamics simulations predicting that neutralization of the charges on these lysine residues would allow more stable parallel in-register packing in this region. We also show empirically that substitution of these clustered lysine residues with alanines or asparagines results in recombinant PrP amyloid fibrils with extended proteinase-K resistant β-sheet cores and infrared spectra that are more reminiscent of bona fide PrP(Sc). These findings indicate that charge neutralization at the central lysine cluster is critical for the folding and tight packing of N-proximal residues within PrP amyloid fibrils. This charge neutralization may be a key aspect of the mechanism by which anionic cofactors promote PrP(Sc) formation.
Highlights
Key factors modulating conversion of prion protein into prions remain unclear
We show empirically that substitution of these clustered lysine residues with alanines or asparagines results in recombinant prion protein (PrP) amyloid fibrils with extended proteinase-K resistant -sheet cores and infrared spectra that are more reminiscent of bona fide PrPSc
These findings indicate that charge neutralization at the central lysine cluster is critical for the folding and tight packing of N-proximal residues within PrP amyloid fibrils
Summary
Key factors modulating conversion of prion protein into prions remain unclear. Results: Neutralization of a cluster of lysines within residues 101–110 promoted formation of an N-terminally extended recombinant prion protein amyloid core. Most pure recombinant prion protein (PrP) amyloids generated in vitro are not infectious and lack the extent of the protease-resistant core and solvent exclusion of infectious PrPSc, especially within residues ϳ90 –160. These findings indicate that charge neutralization at the central lysine cluster is critical for the folding and tight packing of N-proximal residues within PrP amyloid fibrils This charge neutralization may be a key aspect of the mechanism by which anionic cofactors promote PrPSc formation. In brain-derived PrPSc, this region is tightly packed and protease-resistant [6, 7, 22, 23, 27], whereas spontaneously generated noninfectious rPrP fibrils are typically more disordered, solvent-accessible, and protease-sensitive within residues ϳ90 –159 [22, 28].
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