+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-0331 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | Refined 13pf Hela Cell Tubulin microtubule (EML4-NTD decorated) | |||||||||
Map data | Single Asymmetric Unit from symmetrized reconstruction | |||||||||
Sample |
| |||||||||
Function / homology | Function and homology information odontoblast differentiation / Post-chaperonin tubulin folding pathway / Carboxyterminal post-translational modifications of tubulin / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly / cytoskeleton-dependent intracellular transport / Intraflagellar transport / Sealing of the nuclear envelope (NE) by ESCRT-III / Gap junction assembly / Formation of tubulin folding intermediates by CCT/TriC ...odontoblast differentiation / Post-chaperonin tubulin folding pathway / Carboxyterminal post-translational modifications of tubulin / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Cilium Assembly / cytoskeleton-dependent intracellular transport / Intraflagellar transport / Sealing of the nuclear envelope (NE) by ESCRT-III / Gap junction assembly / Formation of tubulin folding intermediates by CCT/TriC / natural killer cell mediated cytotoxicity / COPI-independent Golgi-to-ER retrograde traffic / Kinesins / Assembly and cell surface presentation of NMDA receptors / GTPase activating protein binding / COPI-dependent Golgi-to-ER retrograde traffic / regulation of synapse organization / intercellular bridge / nuclear envelope lumen / Recycling pathway of L1 / RHOH GTPase cycle / spindle assembly / RHO GTPases activate IQGAPs / MHC class I protein binding / Hedgehog 'off' state / cytoplasmic microtubule / microtubule-based process / COPI-mediated anterograde transport / Activation of AMPK downstream of NMDARs / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / Loss of Nlp from mitotic centrosomes / Loss of proteins required for interphase microtubule organization from the centrosome / Recruitment of mitotic centrosome proteins and complexes / Resolution of Sister Chromatid Cohesion / Recruitment of NuMA to mitotic centrosomes / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / Anchoring of the basal body to the plasma membrane / MHC class II antigen presentation / cellular response to interleukin-4 / AURKA Activation by TPX2 / RHO GTPases Activate Formins / Translocation of SLC2A4 (GLUT4) to the plasma membrane / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / PKR-mediated signaling / structural constituent of cytoskeleton / cytoplasmic ribonucleoprotein granule / mitotic spindle / microtubule cytoskeleton organization / Aggrephagy / HCMV Early Events / Separation of Sister Chromatids / The role of GTSE1 in G2/M progression after G2 checkpoint / microtubule cytoskeleton / azurophil granule lumen / Regulation of PLK1 Activity at G2/M Transition / double-stranded RNA binding / mitotic cell cycle / cell body / microtubule / Potential therapeutics for SARS / cytoskeleton / membrane raft / cell division / protein domain specific binding / GTPase activity / ubiquitin protein ligase binding / Neutrophil degranulation / protein-containing complex binding / structural molecule activity / GTP binding / protein-containing complex / extracellular exosome / extracellular region / metal ion binding / nucleus / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) / Human (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.6 Å | |||||||||
Authors | Atherton JM / Moores CA | |||||||||
Funding support | United Kingdom, 2 items
| |||||||||
Citation | Journal: Sci Signal / Year: 2019 Title: Mitotic phosphorylation by NEK6 and NEK7 reduces the microtubule affinity of EML4 to promote chromosome congression. Authors: Rozita Adib / Jessica M Montgomery / Joseph Atherton / Laura O'Regan / Mark W Richards / Kees R Straatman / Daniel Roth / Anne Straube / Richard Bayliss / Carolyn A Moores / Andrew M Fry / Abstract: EML4 is a microtubule-associated protein that promotes microtubule stability. We investigated its regulation across the cell cycle and found that EML4 was distributed as punctate foci along the ...EML4 is a microtubule-associated protein that promotes microtubule stability. We investigated its regulation across the cell cycle and found that EML4 was distributed as punctate foci along the microtubule lattice in interphase but exhibited reduced association with spindle microtubules in mitosis. Microtubule sedimentation and cryo-electron microscopy with 3D reconstruction revealed that the basic N-terminal domain of EML4 mediated its binding to the acidic C-terminal tails of α- and β-tubulin on the microtubule surface. The mitotic kinases NEK6 and NEK7 phosphorylated the EML4 N-terminal domain at Ser and Ser in vitro, and depletion of these kinases in cells led to increased EML4 binding to microtubules in mitosis. An S144A-S146A double mutant not only bound inappropriately to mitotic microtubules but also increased their stability and interfered with chromosome congression. In addition, constitutive activation of NEK6 or NEK7 reduced the association of EML4 with interphase microtubules. Together, these data support a model in which NEK6- and NEK7-dependent phosphorylation promotes the dissociation of EML4 from microtubules in mitosis in a manner that is required for efficient chromosome congression. | |||||||||
History |
|
-Structure visualization
Movie |
Movie viewer |
---|---|
Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_0331.map.gz | 323.6 KB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-0331-v30.xml emd-0331.xml | 15.3 KB 15.3 KB | Display Display | EMDB header |
Images | emd_0331.png | 124.5 KB | ||
Others | emd_0331_additional.map.gz | 282.4 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-0331 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-0331 | HTTPS FTP |
-Related structure data
Related structure data | 6i2iMC M: atomic model generated by this map C: citing same article (ref.) |
---|---|
Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|---|
Related items in Molecule of the Month |
-Map
File | Download / File: emd_0331.map.gz / Format: CCP4 / Size: 307.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Single Asymmetric Unit from symmetrized reconstruction | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.37 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
|
-Supplemental data
-Additional map: 20 angstrom low pass filtered C1 reconstruction
File | emd_0331_additional.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | 20 angstrom low pass filtered C1 reconstruction | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Sample components
-Entire : Alpha and beta-tubulin from Hela Cell (modelled) decorated with E...
Entire | Name: Alpha and beta-tubulin from Hela Cell (modelled) decorated with EML4-NTD (not modelled) |
---|---|
Components |
|
-Supramolecule #1: Alpha and beta-tubulin from Hela Cell (modelled) decorated with E...
Supramolecule | Name: Alpha and beta-tubulin from Hela Cell (modelled) decorated with EML4-NTD (not modelled) type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2 Details: EML4-NTD density at low resolution due to flexibility, thus was not modelled |
---|---|
Source (natural) | Organism: Homo sapiens (human) / Cell: Hela |
Molecular weight | Theoretical: 110 kDa/nm |
-Macromolecule #1: Tubulin alpha-1B chain
Macromolecule | Name: Tubulin alpha-1B chain / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
---|---|
Source (natural) | Organism: Human (human) |
Molecular weight | Theoretical: 50.204445 KDa |
Sequence | String: MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIGGGDDSFN TFFSETGAGK HVPRAVFVDL EPTVIDEVRT GTYRQLFHP EQLITGKEDA ANNYARGHYT IGKEIIDLVL DRIRKLADQC TGLQGFLVFH SFGGGTGSGF TSLLMERLSV D YGKKSKLE ...String: MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIGGGDDSFN TFFSETGAGK HVPRAVFVDL EPTVIDEVRT GTYRQLFHP EQLITGKEDA ANNYARGHYT IGKEIIDLVL DRIRKLADQC TGLQGFLVFH SFGGGTGSGF TSLLMERLSV D YGKKSKLE FSIYPAPQVS TAVVEPYNSI LTTHTTLEHS DCAFMVDNEA IYDICRRNLD IERPTYTNLN RLISQIVSSI TA SLRFDGA LNVDLTEFQT NLVPYPRIHF PLATYAPVIS AEKAYHEQLS VAEITNACFE PANQMVKCDP RHGKYMACCL LYR GDVVPK DVNAAIATIK TKRSIQFVDW CPTGFKVGIN YQPPTVVPGG DLAKVQRAVC MLSNTTAIAE AWARLDHKFD LMYA KRAFV HWYVGEGMEE GEFSEAREDM AALEKDYEEV GVDSVEGEGE EEGEEY |
-Macromolecule #2: Tubulin beta chain
Macromolecule | Name: Tubulin beta chain / type: protein_or_peptide / ID: 2 / Details: Beta1-tubulin / Number of copies: 1 / Enantiomer: LEVO |
---|---|
Source (natural) | Organism: Human (human) |
Molecular weight | Theoretical: 49.717629 KDa |
Sequence | String: MREIVHIQAG QCGNQIGAKF WEVISDEHGI DPTGTYHGDS DLQLDRISVY YNEATGGKYV PRAILVDLEP GTMDSVRSGP FGQIFRPDN FVFGQSGAGN NWAKGHYTEG AELVDSVLDV VRKEAESCDC LQGFQLTHSL GGGTGSGMGT LLISKIREEY P DRIMNTFS ...String: MREIVHIQAG QCGNQIGAKF WEVISDEHGI DPTGTYHGDS DLQLDRISVY YNEATGGKYV PRAILVDLEP GTMDSVRSGP FGQIFRPDN FVFGQSGAGN NWAKGHYTEG AELVDSVLDV VRKEAESCDC LQGFQLTHSL GGGTGSGMGT LLISKIREEY P DRIMNTFS VVPSPKVSDT VVEPYNATLS VHQLVENTDE TYCIDNEALY DICFRTLKLT TPTYGDLNHL VSATMSGVTT CL RFPGQLN ADLRKLAVNM VPFPRLHFFM PGFAPLTSRG SQQYRALTVP ELTQQVFDAK NMMAACDPRH GRYLTVAAVF RGR MSMKEV DEQMLNVQNK NSSYFVEWIP NNVKTAVCDI PPRGLKMAVT FIGNSTAIQE LFKRISEQFT AMFRRKAFLH WYTG EGMDE MEFTEAESNM NDLVSEYQQY QDATAEEEED FGEEAEEEA |
-Macromolecule #3: GUANOSINE-5'-TRIPHOSPHATE
Macromolecule | Name: GUANOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 1 / Formula: GTP |
---|---|
Molecular weight | Theoretical: 523.18 Da |
Chemical component information | ChemComp-GTP: |
-Macromolecule #4: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 2 / Formula: MG |
---|---|
Molecular weight | Theoretical: 24.305 Da |
-Macromolecule #5: PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER
Macromolecule | Name: PHOSPHOMETHYLPHOSPHONIC ACID GUANYLATE ESTER / type: ligand / ID: 5 / Number of copies: 1 / Formula: G2P |
---|---|
Molecular weight | Theoretical: 521.208 Da |
Chemical component information | ChemComp-G2P: |
-Macromolecule #6: TAXOL
Macromolecule | Name: TAXOL / type: ligand / ID: 6 / Number of copies: 1 / Formula: TA1 |
---|---|
Molecular weight | Theoretical: 853.906 Da |
Chemical component information | ChemComp-TA1: |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
Processing | single particle reconstruction |
Aggregation state | filament |
-Sample preparation
Concentration | 0.5 mg/mL |
---|---|
Buffer | pH: 6.8 / Component - Name: BRB25 Details: 25mM PIPES, 1.5mM MgCl2, 1mM EGTA, 1mM DTT, 30mM NaCl,1mM GMPCPP |
Vitrification | Cryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV |
Details | Microtubules formed from Hela cell tubulin |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
---|---|
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 48.0 e/Å2 / Details: Dose-weighted sums used in reconstruction |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Startup model | Type of model: INSILICO MODEL In silico model: Insilico density map of GMPCPP microtubule made from tubulin PDBs |
---|---|
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 19542 |