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Open data
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Basic information
| Entry | Database: PDB / ID: 8to0 | ||||||||||||
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| Title | 48-nm repeating structure of doublets from mouse sperm flagella | ||||||||||||
Components |
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Keywords | STRUCTURAL PROTEIN / Mammalian sperm / axoneme / microtubule-based structure / microtubule inner protein / non-motor proteins / cellular motility / fertility | ||||||||||||
| Function / homology | Function and homology informationERKs are inactivated / receptor signaling protein tyrosine kinase inhibitor activity / protein localization to motile cilium / left/right pattern formation / axonemal microtubule doublet inner sheath / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly / Sealing of the nuclear envelope (NE) by ESCRT-III / outer acrosomal membrane / epithelial cilium movement involved in determination of left/right asymmetry ...ERKs are inactivated / receptor signaling protein tyrosine kinase inhibitor activity / protein localization to motile cilium / left/right pattern formation / axonemal microtubule doublet inner sheath / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly / Sealing of the nuclear envelope (NE) by ESCRT-III / outer acrosomal membrane / epithelial cilium movement involved in determination of left/right asymmetry / regulation of brood size / establishment of left/right asymmetry / 9+0 motile cilium / sperm flagellum assembly / manchette assembly / axonemal B tubule inner sheath / axonemal A tubule inner sheath / Carboxyterminal post-translational modifications of tubulin / Intraflagellar transport / protein polyglutamylation / inner dynein arm assembly / regulation of calcineurin-NFAT signaling cascade / sperm axoneme assembly / regulation of microtubule nucleation / positive regulation of feeding behavior / COPI-independent Golgi-to-ER retrograde traffic / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / Transferases; Transferring phosphorus-containing groups / sperm principal piece / regulation of cilium beat frequency involved in ciliary motility / cerebrospinal fluid circulation / protein tyrosine/serine/threonine phosphatase activity / epithelial cilium movement involved in extracellular fluid movement / cilium movement involved in cell motility / COPI-mediated anterograde transport / Aggrephagy / Kinesins / 9+2 motile cilium / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / regulation of store-operated calcium entry / Resolution of Sister Chromatid Cohesion / intraciliary transport / PKR-mediated signaling / acrosomal membrane / The role of GTSE1 in G2/M progression after G2 checkpoint / RHO GTPases activate IQGAPs / Recycling pathway of L1 / axoneme assembly / microtubule sliding / cilium movement / ciliary transition zone / left/right axis specification / calcium ion sensor activity / axonemal microtubule / COPI-dependent Golgi-to-ER retrograde traffic / RHO GTPases Activate Formins / Separation of Sister Chromatids / organelle transport along microtubule / cilium organization / Hedgehog 'off' state / Loss of Nlp from mitotic centrosomes / Recruitment of mitotic centrosome proteins and complexes / Loss of proteins required for interphase microtubule organization from the centrosome / Recruitment of NuMA to mitotic centrosomes / Anchoring of the basal body to the plasma membrane / forebrain morphogenesis / AURKA Activation by TPX2 / gamma-tubulin ring complex / Regulation of PLK1 Activity at G2/M Transition / manchette / cerebellar cortex morphogenesis / glial cell differentiation / dentate gyrus development / positive regulation of cilium assembly / neuron projection arborization / flagellated sperm motility / MHC class II antigen presentation / cell projection organization / UTP biosynthetic process / CTP biosynthetic process / determination of left/right symmetry / response to L-glutamate / pyramidal neuron differentiation / intermediate filament / centrosome cycle / positive regulation of cell motility / nucleoside diphosphate kinase activity / regulation of focal adhesion assembly / GTP biosynthetic process / tubulin complex / smoothened signaling pathway / AMP binding / protein-serine/threonine phosphatase / ciliary base / regulation of synapse organization / receptor clustering / startle response / negative regulation of epidermal growth factor receptor signaling pathway / adult behavior Similarity search - Function | ||||||||||||
| Biological species | ![]() | ||||||||||||
| Method | ELECTRON MICROSCOPY / subtomogram averaging / cryo EM / Resolution: 7.7 Å | ||||||||||||
Authors | Chen, Z. / Shiozak, M. / Hass, K.M. / Skinner, W. / Zhao, S. / Guo, C. / Polacco, B.J. / Yu, Z. / Krogan, N.J. / Kaake, R.M. ...Chen, Z. / Shiozak, M. / Hass, K.M. / Skinner, W. / Zhao, S. / Guo, C. / Polacco, B.J. / Yu, Z. / Krogan, N.J. / Kaake, R.M. / Vale, R.D. / Agard, D.A. | ||||||||||||
| Funding support | United States, 3items
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Citation | Journal: Cell / Year: 2023Title: De novo protein identification in mammalian sperm using in situ cryoelectron tomography and AlphaFold2 docking. Authors: Zhen Chen / Momoko Shiozaki / Kelsey M Haas / Will M Skinner / Shumei Zhao / Caiying Guo / Benjamin J Polacco / Zhiheng Yu / Nevan J Krogan / Polina V Lishko / Robyn M Kaake / Ronald D Vale / David A Agard / ![]() Abstract: To understand the molecular mechanisms of cellular pathways, contemporary workflows typically require multiple techniques to identify proteins, track their localization, and determine their ...To understand the molecular mechanisms of cellular pathways, contemporary workflows typically require multiple techniques to identify proteins, track their localization, and determine their structures in vitro. Here, we combined cellular cryoelectron tomography (cryo-ET) and AlphaFold2 modeling to address these questions and understand how mammalian sperm are built in situ. Our cellular cryo-ET and subtomogram averaging provided 6.0-Å reconstructions of axonemal microtubule structures. The well-resolved tertiary structures allowed us to unbiasedly match sperm-specific densities with 21,615 AlphaFold2-predicted protein models of the mouse proteome. We identified Tektin 5, CCDC105, and SPACA9 as novel microtubule-associated proteins. These proteins form an extensive interaction network crosslinking the lumen of axonemal doublet microtubules, suggesting their roles in modulating the mechanical properties of the filaments. Indeed, Tekt5 -/- sperm possess more deformed flagella with 180° bends. Together, our studies presented a cellular visual proteomics workflow and shed light on the in vivo functions of Tektin 5. | ||||||||||||
| History |
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Structure visualization
| Structure viewer | Molecule: Molmil Jmol/JSmol |
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Downloads & links
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Download
| PDBx/mmCIF format | 8to0.cif.gz | 20.2 MB | Display | PDBx/mmCIF format |
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| PDB format | pdb8to0.ent.gz | Display | PDB format | |
| PDBx/mmJSON format | 8to0.json.gz | Tree view | PDBx/mmJSON format | |
| Others | Other downloads |
-Validation report
| Summary document | 8to0_validation.pdf.gz | 4.6 MB | Display | wwPDB validaton report |
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| Full document | 8to0_full_validation.pdf.gz | 4.6 MB | Display | |
| Data in XML | 8to0_validation.xml.gz | 2.3 MB | Display | |
| Data in CIF | 8to0_validation.cif.gz | 4.5 MB | Display | |
| Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/to/8to0 ftp://data.pdbj.org/pub/pdb/validation_reports/to/8to0 | HTTPS FTP |
-Related structure data
| Related structure data | ![]() 41431MC C: citing same article ( M: map data used to model this data |
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| Similar structure data | Similarity search - Function & homology F&H Search |
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Links
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Assembly
| Deposited unit | ![]()
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Components
-Cilia- and flagella-associated protein ... , 9 types, 27 molecules 0734AB89AyBHFhFiFjFkFlFmFnBAAcAxdBGBaBoEHEWEl
| #1: Protein | Mass: 26633.035 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) ![]() #10: Protein | Mass: 62036.609 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) ![]() #12: Protein | | Mass: 12278.145 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() #17: Protein | Mass: 23062.510 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) ![]() #18: Protein | Mass: 34433.383 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) ![]() #22: Protein | Mass: 22781.389 Da / Num. of mol.: 7 / Source method: isolated from a natural source / Source: (natural) ![]() #27: Protein | Mass: 65962.016 Da / Num. of mol.: 5 / Source method: isolated from a natural source / Source: (natural) ![]() #28: Protein | Mass: 68322.164 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Source: (natural) ![]() #32: Protein | Mass: 18960.092 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Source: (natural) ![]() |
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+Protein , 22 types, 436 molecules LMIJKNPRTVXZnprtvxzBQBSBUBWBYAWAYAjAlAnAp...
-EF-hand domain-containing family member ... , 2 types, 6 molecules 12FgFSFoFv
| #6: Protein | Mass: 95891.961 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) ![]() #21: Protein | Mass: 87758.023 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) ![]() |
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-Piercer of microtubule wall ... , 2 types, 2 molecules GFw
| #25: Protein | Mass: 13728.513 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() |
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| #35: Protein | Mass: 18862.852 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() |
-Experimental details
-Experiment
| Experiment | Method: ELECTRON MICROSCOPY |
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| EM experiment | Aggregation state: CELL / 3D reconstruction method: subtomogram averaging |
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Sample preparation
| Component | Name: Mouse sperm / Type: CELL / Entity ID: all / Source: NATURAL |
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| Source (natural) | Organism: ![]() |
| Buffer solution | pH: 7.4 |
| Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
| Vitrification | Cryogen name: ETHANE |
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Electron microscopy imaging
| Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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| Microscopy | Model: FEI TITAN KRIOS |
| Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
| Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 6000 nm / Nominal defocus min: 2000 nm |
| Image recording | Electron dose: 4 e/Å2 / Avg electron dose per subtomogram: 100 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
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Processing
| EM software | Name: RELION / Version: 4.0-beta2 / Category: 3D reconstruction |
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| CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION |
| Symmetry | Point symmetry: C1 (asymmetric) |
| 3D reconstruction | Resolution: 7.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 12848 / Symmetry type: POINT |
| EM volume selection | Details: 32288 particles were initially picked every 24 nm along the microtubules. Num. of tomograms: 76 / Num. of volumes extracted: 32288 |
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FIELD EMISSION GUN