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Yorodumi- EMDB-7005: Negative stain reconstruction of the peroxisomal AAA-ATPase Pex1/... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-7005 | ||||||||||||
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Title | Negative stain reconstruction of the peroxisomal AAA-ATPase Pex1/Pex6 complex associated with substrate Pex15 | ||||||||||||
Map data | Negative stain reconstruction of the Pex15 bound Pex1-Pex6 complex. | ||||||||||||
Sample |
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Biological species | Saccharomyces cerevisiae (brewer's yeast) | ||||||||||||
Method | single particle reconstruction / negative staining / Resolution: 23.2 Å | ||||||||||||
Authors | Chowdhury S / Gardner BM / Castanzo DT / Stjepanovic G / Stefely MS / Hurley JH / Martin A / Lander GC | ||||||||||||
Funding support | United States, 3 items
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Citation | Journal: Nat Commun / Year: 2018 Title: The peroxisomal AAA-ATPase Pex1/Pex6 unfolds substrates by processive threading. Authors: Brooke M Gardner / Dominic T Castanzo / Saikat Chowdhury / Goran Stjepanovic / Matthew S Stefely / James H Hurley / Gabriel C Lander / Andreas Martin / Abstract: Pex1 and Pex6 form a heterohexameric motor essential for peroxisome biogenesis and function, and mutations in these AAA-ATPases cause most peroxisome-biogenesis disorders in humans. The tail-anchored ...Pex1 and Pex6 form a heterohexameric motor essential for peroxisome biogenesis and function, and mutations in these AAA-ATPases cause most peroxisome-biogenesis disorders in humans. The tail-anchored protein Pex15 recruits Pex1/Pex6 to the peroxisomal membrane, where it performs an unknown function required for matrix-protein import. Here we determine that Pex1/Pex6 from S. cerevisiae is a protein translocase that unfolds Pex15 in a pore-loop-dependent and ATP-hydrolysis-dependent manner. Our structural studies of Pex15 in isolation and in complex with Pex1/Pex6 illustrate that Pex15 binds the N-terminal domains of Pex6, before its C-terminal disordered region engages with the pore loops of the motor, which then processively threads Pex15 through the central pore. Furthermore, Pex15 directly binds the cargo receptor Pex5, linking Pex1/Pex6 to other components of the peroxisomal import machinery. Our results thus support a role of Pex1/Pex6 in mechanical unfolding of peroxins or their extraction from the peroxisomal membrane during matrix-protein import. | ||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_7005.map.gz | 1.8 MB | EMDB map data format | |
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Header (meta data) | emd-7005-v30.xml emd-7005.xml | 12.5 KB 12.5 KB | Display Display | EMDB header |
Images | emd_7005.png | 58.5 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-7005 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-7005 | HTTPS FTP |
-Validation report
Summary document | emd_7005_validation.pdf.gz | 78.8 KB | Display | EMDB validaton report |
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Full document | emd_7005_full_validation.pdf.gz | 77.9 KB | Display | |
Data in XML | emd_7005_validation.xml.gz | 493 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-7005 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-7005 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_7005.map.gz / Format: CCP4 / Size: 2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Negative stain reconstruction of the Pex15 bound Pex1-Pex6 complex. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 4.1 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Complex between Pex1-Pex6 AAA-ATPase with substrate Pex15
Entire | Name: Complex between Pex1-Pex6 AAA-ATPase with substrate Pex15 |
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Components |
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-Supramolecule #1: Complex between Pex1-Pex6 AAA-ATPase with substrate Pex15
Supramolecule | Name: Complex between Pex1-Pex6 AAA-ATPase with substrate Pex15 type: complex / ID: 1 / Parent: 0 |
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Source (natural) | Organism: Saccharomyces cerevisiae (brewer's yeast) |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant strain: BL21 |
Molecular weight | Theoretical: 750 KDa |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.6 Details: 60 mM HEPES pH 7.6, 50 mM NaCl, 50 mM KCl, 10 % glycerol, 10 mM MgCl2, 0.5 mM EDTA and 5mM ATP |
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Staining | Type: NEGATIVE / Material: Uranyl Formate |
Grid | Model: Maxtaform / Material: COPPER/RHODIUM / Mesh: 400 / Support film - Material: CARBON / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR |
-Electron microscopy
Microscope | FEI TECNAI SPIRIT |
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Temperature | Max: 298.15 K |
Image recording | Film or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Digitization - Sampling interval: 15.6 µm / Number grids imaged: 1 / Number real images: 1097 / Average exposure time: 0.4 sec. / Average electron dose: 20.0 e/Å2 |
Electron beam | Acceleration voltage: 120 kV / Electron source: LAB6 |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.2 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 52000 |
Sample stage | Specimen holder model: SIDE ENTRY, EUCENTRIC |
Experimental equipment | Model: Tecnai Spirit / Image courtesy: FEI Company |