Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2022 Jul 13;13(1):4004.
doi: 10.1038/s41467-022-30457-7.

2.7 Å cryo-EM structure of ex vivo RML prion fibrils

Szymon W Manka  1 , Wenjuan Zhang  1 , Adam Wenborn  1 , Jemma Betts  1 , Susan Joiner  1 , Helen R Saibil  2 , John Collinge  3 , Jonathan D F Wadsworth  4
Affiliations

2.7 Å cryo-EM structure of ex vivo RML prion fibrils

Szymon W Manka et al. Nat Commun. .

Abstract

Mammalian prions propagate as distinct strains and are composed of multichain assemblies of misfolded host-encoded prion protein (PrP). Here, we present a near-atomic resolution cryo-EM structure of PrP fibrils present in highly infectious prion rod preparations isolated from the brains of RML prion-infected mice. We found that prion rods comprise single-protofilament helical amyloid fibrils that coexist with twisted pairs of the same protofilaments. Each rung of the protofilament is formed by a single PrP monomer with the ordered core comprising PrP residues 94-225, which folds to create two asymmetric lobes with the N-linked glycans and the glycosylphosphatidylinositol anchor projecting from the C-terminal lobe. The overall architecture is comparable to that of recently reported PrP fibrils isolated from the brain of hamsters infected with the 263K prion strain. However, there are marked conformational variations that could result from differences in PrP sequence and/or represent distinguishing features of the distinct prion strains.

PubMed Disclaimer

Conflict of interest statement

J.C. is a Director and J.C. and J.D.F.W. are shareholders of D-Gen Limited, an academic spin-out company working in the field of prion disease diagnosis, decontamination, and therapeutics. D-Gen supplied the ICSM35 and ICSM18 antibodies used for western blot and ELISA performed in this study. The other authors declare no competing interests.

Figures

Fig. 1
Fig. 1. RML fibril morphologies and 3D reconstruction and atomic model of the RML protofilament core.
a Selected cryo-EM images (300 kV FEI Krios G3i, K3 camera) showing examples of single RML protofilaments (s) and paired protofilaments (p) with approximate measurements of widths and crossover distances. Black arrowheads, partly intertwined protofilaments; white arrowheads, ‘notched’ paired protofilaments. Pie chart, quantification of paired protofilaments in 6000 micrographs. b Rendered cryo-EM map (isosurface) view of both sides of a helical crossover, with annotated locations of N-linked glycans and the GPI-anchor. c Cross-section (as indicated in b) of the unrendered map (pixel size: 1.067 Å) showing the protein core and the non-protein extra densities, with annotations coloured as in b; *, likely locations of phosphotungstate polyanions (cages) near positively charged residues;?, unassigned density. See also Supplementary Fig. 4b. d Protein-only density of a single amyloid rung with the fitted atomic model of the mouse PrP chain shown with sticks and coloured by heteroatom: C, green; N, blue; O, red; S, yellow. e Local resolution of the map calculated with Relion 3.1 LocRes. *, as in c. f Diagram of the PrP subunit. Positions of amino acid side chains are indicated with circles (positively charged, blue; negatively charged, red; neutral, green; hydrophobic, white; aromatic, grey) on either side of the backbone (black line). β-strands are indicated with thick black arrow-headed lines.
Fig. 2
Fig. 2. Intra- and inter-chain interactions stabilising the RML prion fibril.
a Schematic depiction of the alternating polar and non-polar lateral contacts that stabilise a single PrP monomer in the fibril. Transparent surface representation is coloured by hydrophobicity (hydrophobic, yellow; hydrophilic, teal). Protein backbone is shown with cartoon (licorice) representation and amino acid side chains as white sticks coloured by heteroatom (O, red; N, blue; S, yellow). b Butterfly view of the top and bottom surface of each rung, coloured as in a. Dotted lines indicate longitudinally connecting regions in the assembly. c Butterfly view of charge distribution in the assembly. Dotted lines as in b. d Ribbon representation of 3 amyloid rungs, with indicated β-sheets and inter-chain hydrogen bonds (dotted lines).
Fig. 3
Fig. 3. Staggering of the N- and C-terminal lobes in the RML prion fibril.
Top, cryo-EM density (MAP) and solvent-excluded surface (MODEL) of three rungs, with indicated side chains that connect to form the inter-lobe contacts. The F174 and H176 residues are better visible in the MAP and MODEL views, respectively. Bottom, magnified view of the staggered interactions (hydrophobic contacts) shown with transparent surface, main chain as ribbon and selected interacting residues as sticks.
Fig. 4
Fig. 4. Comparison of PrP conformation in the RML and the 263K prion fibrils.
a Polypeptide backbone superposition on the first two β-strands (N-term alignment; secondary structure not shown) of single PrP monomers from the two different strains, coloured by their deviation in distance. b Surface models showing internal gaps and the divergent angles between the N- and C-terminal lobes. c Top, multiple PrP sequence alignment coloured by conservation and annotated by mouse RML PrP sequence numbering and secondary structure. Mouse vs hamster amino acid substitutions (AAS) that underpin distinct conformations of RML and 263K fibrils are highlighted in red. Bottom, mapping of the selected AAS onto cartoon structures of RML (this study) and 263K (PDB code: 7LNA) fibrils (sticks coloured white and by heteroatom: O, red; S, yellow). Selected conserved residues, including those involved in distinct interactions due to divergent PrP folds are shown with sticks coloured as main chain and by heteroatom (N, blue; O, red; S, yellow). Red arrows indicate different folds of the C-termini, which result in divergent tips of the C-terminal lobes.
See this image and copyright information in PMC

Comment in

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Prusiner SB. Prions. Proc. Natl Acad. Sci. USA. 1998;95:13363–13383. doi: 10.1073/pnas.95.23.13363. - DOI - PMC - PubMed
    1. Collinge J, Clarke A. A general model of prion strains and their pathogenicity. Science. 2007;318:930–936. doi: 10.1126/science.1138718. - DOI - PubMed
    1. Collinge J. Mammalian prions and their wider relevance in neurodegenerative diseases. Nature. 2016;539:217–226. doi: 10.1038/nature20415. - DOI - PubMed
    1. Terry C, Wadsworth JDF. Recent advances in understanding mammalian prion structure: a mini review. Front Mol. Neurosci. 2019;12:169. doi: 10.3389/fnmol.2019.00169. - DOI - PMC - PubMed
    1. Goedert M. Neurodegeneration. Alzheimer’s and Parkinson’s diseases: the prion concept in relation to assembled Abeta, tau, and alpha-synuclein. Science. 2015;349:1255555. doi: 10.1126/science.1255555. - DOI - PubMed

Publication types