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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-2946 | |||||||||
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Title | cryo-EM structure of HET-s prion infectious form | |||||||||
![]() | Reconstruction of HET-s prion amyloid assembled at pH3 | |||||||||
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![]() | amyloid / prion / cross-beta structure | |||||||||
Function / homology | Het-s prion-forming domain / Het-s prion-forming domain / Prion-inhibition and propagation, HeLo domain / HeLo domain superfamily / Het-s 218-289 / Prion-inhibition and propagation / identical protein binding / cytoplasm / Heterokaryon incompatibility protein s![]() | |||||||||
Biological species | ![]() | |||||||||
Method | helical reconstruction / cryo EM / Resolution: 8.5 Å | |||||||||
![]() | Mizuno N / Baxa U / Steven AC | |||||||||
![]() | ![]() Title: Structural dependence of HET-s amyloid fibril infectivity assessed by cryoelectron microscopy. Authors: Naoko Mizuno / Ulrich Baxa / Alasdair C Steven / ![]() Abstract: HET-s is a prion protein of the fungus Podospora anserina which, in the prion state, is active in a self/nonself recognition process called heterokaryon incompatibility. Its prionogenic properties ...HET-s is a prion protein of the fungus Podospora anserina which, in the prion state, is active in a self/nonself recognition process called heterokaryon incompatibility. Its prionogenic properties reside in the C-terminal "prion domain." The HET-s prion domain polymerizes in vitro into amyloid fibrils whose properties depend on the pH of assembly; above pH 3, infectious singlet fibrils are produced, and below pH 3, noninfectious triplet fibrils. To investigate the correlation between structure and infectivity, we performed cryo-EM analyses. Singlet fibrils have a helical pitch of approximately 410 Å and a left-handed twist. Triplet fibrils have three protofibrils whose lateral dimensions (36 × 25 Å) and axial packing (one subunit per 9.4 Å) match those of singlets but differ in their supercoiling. At 8.5-Å resolution, the cross-section of the singlet fibril reconstruction is largely consistent with that of a β-solenoid model previously determined by solid-state NMR. Reconstructions of the triplet fibrils show three protofibrils coiling around a common axis and packed less tightly at pH 3 than at pH 2, eventually peeling off. Taken together with the earlier observation that fragmentation of triplet fibrils by sonication does not increase infectivity, these observations suggest a novel mechanism for self-propagation, whereby daughter fibrils nucleate on the lateral surface of singlet fibrils. In triplets, this surface is occluded, blocking nucleation and thereby explaining their lack of infectivity. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 548.5 KB | ![]() | |
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Header (meta data) | ![]() ![]() | 10 KB 10 KB | Display Display | ![]() |
Images | ![]() | 62.9 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 173.5 KB | Display | ![]() |
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Full document | ![]() | 172.6 KB | Display | |
Data in XML | ![]() | 4.1 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Reconstruction of HET-s prion amyloid assembled at pH3 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.84 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : HET-s prion domain (218-289) assembled at pH3
Entire | Name: HET-s prion domain (218-289) assembled at pH3 |
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Components |
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-Supramolecule #1000: HET-s prion domain (218-289) assembled at pH3
Supramolecule | Name: HET-s prion domain (218-289) assembled at pH3 / type: sample / ID: 1000 Oligomeric state: helical polymer with a rise of 9.4 A per protein unit Number unique components: 1 |
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Molecular weight | Experimental: 8 KDa |
-Macromolecule #1: Heterokaryon incompatibility protein s
Macromolecule | Name: Heterokaryon incompatibility protein s / type: protein_or_peptide / ID: 1 / Name.synonym: HET-s Details: HET-s proteins are assembled to make a single strand helix, with a rise of 9.4 A per protein. Oligomeric state: Helical filament / Recombinant expression: Yes |
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Source (natural) | Organism: ![]() |
Molecular weight | Experimental: 8 KDa / Theoretical: 8 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | UniProtKB: Heterokaryon incompatibility protein s / InterPro: Het-s prion-forming domain |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | helical reconstruction |
Aggregation state | filament |
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Sample preparation
Concentration | 1 mg/mL |
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Buffer | pH: 3 / Details: 20 mM citrate acid |
Grid | Details: Glow discharged Quantifoil R2/2 was used. |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 90 % / Instrument: FEI VITROBOT MARK II / Method: Blot for 5 seconds before plunging |
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Electron microscopy
Microscope | FEI/PHILIPS CM200FEG |
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Temperature | Min: 90 K / Max: 100 K |
Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 120,000 times magnification |
Date | Apr 1, 2008 |
Image recording | Category: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Digitization - Sampling interval: 7 µm / Number real images: 100 / Average electron dose: 20 e/Å2 / Bits/pixel: 8 |
Electron beam | Acceleration voltage: 120 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 38000 |
Sample stage | Specimen holder model: GATAN LIQUID NITROGEN |
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Image processing
Details | The reconstruction was performed using SPIDER in combination with IHRSR. |
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Final reconstruction | Applied symmetry - Helical parameters - Δz: 18.8 Å Applied symmetry - Helical parameters - Δ&Phi: 17.1 ° Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric) Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 8.5 Å / Resolution method: OTHER / Software - Name: SPIder, EMAN, IHRSR, bsoft |
CTF correction | Details: Each micrograph |