+Open data
-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 information ERKs are inactivated / 9+0 motile cilium / sperm flagellum assembly / outer acrosomal membrane / regulation of brood size / establishment of left/right asymmetry / protein localization to motile cilium / manchette assembly / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly ...ERKs are inactivated / 9+0 motile cilium / sperm flagellum assembly / outer acrosomal membrane / regulation of brood size / establishment of left/right asymmetry / protein localization to motile cilium / manchette assembly / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly / Sealing of the nuclear envelope (NE) by ESCRT-III / left/right pattern formation / axonemal B tubule inner sheath / axonemal A tubule inner sheath / epithelial cilium movement involved in determination of left/right asymmetry / regulation of calcineurin-NFAT signaling cascade / MAP kinase phosphatase activity / inner dynein arm assembly / Intraflagellar transport / Carboxyterminal post-translational modifications of tubulin / protein polyglutamylation / positive regulation of feeding behavior / cerebrospinal fluid circulation / sperm axoneme assembly / COPI-independent Golgi-to-ER retrograde traffic / cilium-dependent cell motility / sperm principal piece / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / regulation of cilium beat frequency involved in ciliary motility / cilium movement involved in cell motility / regulation of store-operated calcium entry / epithelial cilium movement involved in extracellular fluid movement / 9+2 motile cilium / intraciliary transport / PKR-mediated signaling / negative regulation of chemotaxis / COPI-mediated anterograde transport / calcium ion sensor activity / Aggrephagy / Transferases; Transferring phosphorus-containing groups / acrosomal membrane / Kinesins / ciliary transition zone / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / cilium movement / Resolution of Sister Chromatid Cohesion / RHO GTPases activate IQGAPs / The role of GTSE1 in G2/M progression after G2 checkpoint / Recycling pathway of L1 / axoneme assembly / axonemal microtubule / negative regulation of T cell activation / left/right axis specification / cilium organization / COPI-dependent Golgi-to-ER retrograde traffic / gamma-tubulin ring complex / flagellated sperm motility / RHO GTPases Activate Formins / Separation of Sister Chromatids / Hedgehog 'off' state / organelle transport along microtubule / glial cell differentiation / Loss of Nlp from mitotic centrosomes / Recruitment of mitotic centrosome proteins and complexes / Loss of proteins required for interphase microtubule organization from the centrosome / protein tyrosine/serine/threonine phosphatase activity / Recruitment of NuMA to mitotic centrosomes / Anchoring of the basal body to the plasma membrane / AURKA Activation by TPX2 / forebrain morphogenesis / neuron projection arborization / Regulation of PLK1 Activity at G2/M Transition / positive regulation of focal adhesion disassembly / manchette / cerebellar cortex morphogenesis / positive regulation of cilium assembly / dentate gyrus development / MHC class II antigen presentation / pyramidal neuron differentiation / negative regulation of JNK cascade / CTP biosynthetic process / UTP biosynthetic process / motile cilium / positive regulation of cell motility / determination of left/right symmetry / centrosome cycle / motor behavior / GTP biosynthetic process / microtubule organizing center / response to L-glutamate / intermediate filament / regulation of neuron projection development / ciliary base / nucleoside diphosphate kinase activity / regulation of focal adhesion assembly / tubulin complex / negative regulation of T cell receptor signaling pathway / negative regulation of MAPK cascade / smoothened signaling pathway Similarity search - Function | ||||||||||||
Biological species | Mus musculus (house mouse) | ||||||||||||
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: 2023 Title: 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: MolmilJmol/JSmol |
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-Downloads & links
-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 (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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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) Mus musculus (house mouse) / References: UniProt: Q9CQC3 #10: Protein | Mass: 62036.609 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q9D439 #12: Protein | | Mass: 12278.145 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q9D9D9 #17: Protein | Mass: 23062.510 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q4KKZ1 #18: Protein | Mass: 34433.383 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q6P8Y0 #22: Protein | Mass: 22781.389 Da / Num. of mol.: 7 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q8BTU1 #27: Protein | Mass: 65962.016 Da / Num. of mol.: 5 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q9D9U9 #28: Protein | Mass: 68322.164 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q5F201 #32: Protein | Mass: 18960.092 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q9DAD0 |
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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) Mus musculus (house mouse) / References: UniProt: Q8CDU5 #21: Protein | Mass: 87758.023 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q9D485 |
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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) Mus musculus (house mouse) / References: UniProt: V9GXK1 |
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#35: Protein | Mass: 18862.852 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Mus musculus (house mouse) / References: UniProt: Q5BN45 |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: CELL / 3D reconstruction method: subtomogram averaging |
-Sample preparation
Component | Name: Mouse sperm / Type: CELL / Entity ID: all / Source: NATURAL |
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Source (natural) | Organism: Mus musculus (house mouse) |
Buffer solution | pH: 7.4 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-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) |
-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 |