+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 / protein localization to motile cilium / outer acrosomal membrane / epithelial cilium movement involved in determination of left/right asymmetry / left/right pattern formation / regulation of brood size / establishment of left/right asymmetry / 9+0 motile cilium / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly ...ERKs are inactivated / protein localization to motile cilium / outer acrosomal membrane / epithelial cilium movement involved in determination of left/right asymmetry / left/right pattern formation / regulation of brood size / establishment of left/right asymmetry / 9+0 motile cilium / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly / Sealing of the nuclear envelope (NE) by ESCRT-III / regulation of calcineurin-NFAT signaling cascade / MAP kinase phosphatase activity / Intraflagellar transport / sperm axoneme assembly / inner dynein arm assembly / cilium-dependent cell motility / Carboxyterminal post-translational modifications of tubulin / protein polyglutamylation / positive regulation of feeding behavior / cerebrospinal fluid circulation / sperm principal piece / COPI-independent Golgi-to-ER retrograde traffic / regulation of cilium beat frequency involved in ciliary motility / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / epithelial cilium movement involved in extracellular fluid movement / cilium movement involved in cell motility / intraciliary transport / negative regulation of chemotaxis / 9+2 motile cilium / regulation of store-operated calcium entry / PKR-mediated signaling / COPI-mediated anterograde transport / cilium movement / Aggrephagy / Transferases; Transferring phosphorus-containing groups / calcium ion sensor activity / acrosomal membrane / Kinesins / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / Resolution of Sister Chromatid Cohesion / RHO GTPases activate IQGAPs / axoneme assembly / The role of GTSE1 in G2/M progression after G2 checkpoint / flagellated sperm motility / cilium organization / Recycling pathway of L1 / negative regulation of T cell activation / axonemal microtubule / left/right axis specification / COPI-dependent Golgi-to-ER retrograde traffic / gamma-tubulin ring complex / 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 / Recruitment of NuMA to mitotic centrosomes / protein tyrosine/serine/threonine phosphatase activity / 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 / manchette / positive regulation of focal adhesion disassembly / positive regulation of cilium assembly / cerebellar cortex morphogenesis / dentate gyrus development / MHC class II antigen presentation / pyramidal neuron differentiation / negative regulation of JNK cascade / UTP biosynthetic process / CTP biosynthetic process / motile cilium / positive regulation of cell motility / determination of left/right symmetry / GTP biosynthetic process / centrosome cycle / intermediate filament / motor behavior / microtubule organizing center / nucleoside diphosphate kinase activity / tubulin complex / response to L-glutamate / ciliary base / negative regulation of T cell receptor signaling pathway / regulation of focal adhesion assembly / smoothened signaling pathway / myosin phosphatase activity / adult behavior / intercellular bridge / regulation of synapse organization / peptidyl-tyrosine dephosphorylation involved in inactivation of protein kinase activity / negative regulation of MAPK cascade / negative regulation of epidermal growth factor receptor 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
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/to/8to0 ftp://data.pdbj.org/pub/pdb/validation_reports/to/8to0 | HTTPS FTP |
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-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 FIELDBright-field microscopy / 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 |