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Yorodumi- EMDB-2943: Structural characterization of the Olfactomedin-1 disulfide-linke... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-2943 | |||||||||
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Title | Structural characterization of the Olfactomedin-1 disulfide-linked tetramer | |||||||||
Map data | Subtomogram of an Olfactomedin-1 tetramer | |||||||||
Sample |
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Keywords | Olfactomedin-1 (Olfm1) / neurobiology / tetramer / X-ray crystallography / small-angle X-ray scattering (SAXS) / electron tomography / analytical ultracentrifugation (AUC) / coiled coil / calcium / disulfide | |||||||||
Biological species | Mus musculus (house mouse) | |||||||||
Method | electron tomography / negative staining | |||||||||
Authors | Sharp TH / Pronker MF / Bos TG / Thies-Weesie DM / Janssen BJC | |||||||||
Citation | Journal: J Biol Chem / Year: 2015 Title: Olfactomedin-1 Has a V-shaped Disulfide-linked Tetrameric Structure. Authors: Matti F Pronker / Trusanne G A A Bos / Thomas H Sharp / Dominique M E Thies-Weesie / Bert J C Janssen / Abstract: Olfactomedin-1 (Olfm1; also known as noelin and pancortin) is a member of the olfactomedin domain-containing superfamily and a highly expressed neuronal glycoprotein important for nervous system ...Olfactomedin-1 (Olfm1; also known as noelin and pancortin) is a member of the olfactomedin domain-containing superfamily and a highly expressed neuronal glycoprotein important for nervous system development. It binds a number of secreted proteins and cell surface-bound receptors to induce cell signaling processes. Using a combined approach of x-ray crystallography, solution scattering, analytical ultracentrifugation, and electron microscopy we determined that full-length Olfm1 forms disulfide-linked tetramers with a distinctive V-shaped architecture. The base of the "V" is formed by two disulfide-linked dimeric N-terminal domains. Each of the two V legs consists of a parallel dimeric disulfide-linked coiled coil with a C-terminal β-propeller dimer at the tips. This agrees with our crystal structure of a C-terminal coiled-coil segment and β-propeller combination (Olfm1(coil-Olf)) that reveals a disulfide-linked dimeric arrangement with the β-propeller top faces in an outward exposed orientation. Similar to its family member myocilin, Olfm1 is stabilized by calcium. The dimer-of-dimers architecture suggests a role for Olfm1 in clustering receptors to regulate signaling and sheds light on the conformation of several other olfactomedin domain family members. | |||||||||
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_2943.map.gz | 405.8 KB | EMDB map data format | |
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Header (meta data) | emd-2943-v30.xml emd-2943.xml | 11.7 KB 11.7 KB | Display Display | EMDB header |
Images | 2943.png | 64.2 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-2943 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-2943 | HTTPS FTP |
-Validation report
Summary document | emd_2943_validation.pdf.gz | 125.8 KB | Display | EMDB validaton report |
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Full document | emd_2943_full_validation.pdf.gz | 125 KB | Display | |
Data in XML | emd_2943_validation.xml.gz | 5.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2943 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2943 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_2943.map.gz / Format: CCP4 / Size: 3.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Subtomogram of an Olfactomedin-1 tetramer | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 4.57 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 : Olfactomedin-1 disulfide-linked tetramer
Entire | Name: Olfactomedin-1 disulfide-linked tetramer |
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Components |
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-Supramolecule #1000: Olfactomedin-1 disulfide-linked tetramer
Supramolecule | Name: Olfactomedin-1 disulfide-linked tetramer / type: sample / ID: 1000 / Oligomeric state: Tetramer / Number unique components: 1 |
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Molecular weight | Experimental: 242 KDa / Theoretical: 256 KDa / Method: AUC |
-Supramolecule #1: Olfactomedin-1
Supramolecule | Name: Olfactomedin-1 / type: organelle_or_cellular_component / ID: 1 / Name.synonym: Olfm1, noelin, pancortin Details: Purified recombinant full-length glycoslated Olfactomedin-1 tetramers were negatively stained with uranyl formate. Number of copies: 1 / Oligomeric state: Tetramer / Recombinant expression: Yes |
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Ref INTERPRO | 0: IPR022082 |
Ref INTERPRO | 0: IPR003112 |
Source (natural) | Organism: Mus musculus (house mouse) / synonym: House Mouse / Tissue: brain / Cell: neuron / Organelle: Secretory pathway / Location in cell: Secreted (ER/golgi/extracellular) |
Molecular weight | Experimental: 242 KDa / Theoretical: 256 KDa |
Recombinant expression | Organism: Homo sapiens (human) / Recombinant cell: HEK293ES / Recombinant plasmid: pUPE107.03 |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | electron tomography |
Aggregation state | particle |
-Sample preparation
Concentration | 0.065 mg/mL |
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Buffer | pH: 7.5 / Details: 150 mM NaCl, 20 mM HEPES |
Staining | Type: NEGATIVE Details: Full-length Olfm1 was adsorbed to grids for 30 sec. Grids were briefly washed with water and then stained for 30 sec with a freshly prepared filtered 2% uranyl formate solution. |
Grid | Details: Carbon-coated mesh copper grids glow discharged for 15 sec |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
-Electron microscopy
Microscope | FEI TECNAI F20 |
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Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected at 30,000 times magnification |
Details | Weak beam illumination |
Date | Feb 12, 2015 |
Image recording | Category: CCD / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k) / Number real images: 58 / Bits/pixel: 32 |
Electron beam | Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 30000 |
Sample stage | Specimen holder model: SIDE ENTRY, EUCENTRIC / Tilt series - Axis1 - Min angle: -58 ° / Tilt series - Axis1 - Max angle: 58 ° / Tilt series - Axis1 - Angle increment: 2 ° |
Experimental equipment | Model: Tecnai F20 / Image courtesy: FEI Company |
-Image processing
Details | Tilt series were collected from -58 to 58 degrees in 2 degree increments. The first and last image was discarded. CTF correction was performed on each projection |
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Final reconstruction | Algorithm: OTHER / Software - Name: IMOD, EMAN2 Details: Sub-tomogram particles were manually picked using e2spt_boxer.py from EMAN2. Each particle was normalized and masked with a sharp spherical mask to remove background density not associated ...Details: Sub-tomogram particles were manually picked using e2spt_boxer.py from EMAN2. Each particle was normalized and masked with a sharp spherical mask to remove background density not associated with the protein. Particles were then filtered to 20 A with a low-pass Gaussian filter, before a tight mask was applied to the remaining density using e2proc3d.py from EMAN2. Number images used: 57 |
CTF correction | Details: IMOD, each tilt image |
-Atomic model buiding 1
Initial model | PDB ID: Chain - #0 - Chain ID: A / Chain - #1 - Chain ID: B |
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Software | Name: Chimera |
Details | Each coiled-coil dimer of the tetramer was fitted as a rigid body. |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |