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Yorodumi- EMDB-15118: 13pf undecorated microtubule from recombinant human tubulin (alph... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-15118 | |||||||||
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Title | 13pf undecorated microtubule from recombinant human tubulin (alpha1B, beta3) lacking the C-terminal tail | |||||||||
Map data | Human microtubules (alpha1B, beta3) lacking the C-terminal tail. | |||||||||
Sample |
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Keywords | tubulin / no tail / recombinant / semisynthetic / STRUCTURAL PROTEIN | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | helical reconstruction / cryo EM / Resolution: 3.68 Å | |||||||||
Authors | Ebberink E / Fernandes S / Hatzopoulos GN / Agashe N / Guidotti N / Reichart T / Reymond L / Velluz MC / Schneider FZ / Pourroy C ...Ebberink E / Fernandes S / Hatzopoulos GN / Agashe N / Guidotti N / Reichart T / Reymond L / Velluz MC / Schneider FZ / Pourroy C / Janke C / Gonczy P / Aumeier C / Fierz B | |||||||||
Funding support | Switzerland, 1 items
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Citation | Journal: Nat Chem / Year: 2023 Title: Tubulin engineering by semi-synthesis reveals that polyglutamylation directs detyrosination. Authors: Eduard Ebberink / Simon Fernandes / Georgios Hatzopoulos / Ninad Agashe / Po-Han Chang / Nora Guidotti / Timothy M Reichart / Luc Reymond / Marie-Claire Velluz / Fabian Schneider / Cédric ...Authors: Eduard Ebberink / Simon Fernandes / Georgios Hatzopoulos / Ninad Agashe / Po-Han Chang / Nora Guidotti / Timothy M Reichart / Luc Reymond / Marie-Claire Velluz / Fabian Schneider / Cédric Pourroy / Carsten Janke / Pierre Gönczy / Beat Fierz / Charlotte Aumeier / Abstract: Microtubules, a critical component of the cytoskeleton, carry post-translational modifications (PTMs) that are important for the regulation of key cellular processes. Long-lived microtubules, in ...Microtubules, a critical component of the cytoskeleton, carry post-translational modifications (PTMs) that are important for the regulation of key cellular processes. Long-lived microtubules, in neurons particularly, exhibit both detyrosination of α-tubulin and polyglutamylation. Dysregulation of these PTMs can result in developmental defects and neurodegeneration. Owing to a lack of tools to study the regulation and function of these PTMs, the mechanisms that govern such PTM patterns are not well understood. Here we produce fully functional tubulin carrying precisely defined PTMs within its C-terminal tail. We ligate synthetic α-tubulin tails-which are site-specifically glutamylated-to recombinant human tubulin heterodimers by applying a sortase- and intein-mediated tandem transamidation strategy. Using microtubules reconstituted with these designer tubulins, we find that α-tubulin polyglutamylation promotes its detyrosination by enhancing the activity of the tubulin tyrosine carboxypeptidase vasohibin/small vasohibin-binding protein in a manner dependent on the length of polyglutamyl chains. We also find that modulating polyglutamylation levels in cells results in corresponding changes in detyrosination, corroborating the link between the detyrosination cycle to polyglutamylation. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_15118.map.gz | 762.9 MB | EMDB map data format | |
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Header (meta data) | emd-15118-v30.xml emd-15118.xml | 16.3 KB 16.3 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_15118_fsc.xml | 19.9 KB | Display | FSC data file |
Images | emd_15118.png | 109.9 KB | ||
Masks | emd_15118_msk_1.map | 824 MB | Mask map | |
Others | emd_15118_half_map_1.map.gz emd_15118_half_map_2.map.gz | 763.4 MB 763.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-15118 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-15118 | HTTPS FTP |
-Validation report
Summary document | emd_15118_validation.pdf.gz | 1.1 MB | Display | EMDB validaton report |
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Full document | emd_15118_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | emd_15118_validation.xml.gz | 29.2 KB | Display | |
Data in CIF | emd_15118_validation.cif.gz | 38.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-15118 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-15118 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_15118.map.gz / Format: CCP4 / Size: 824 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Annotation | Human microtubules (alpha1B, beta3) lacking the C-terminal tail. | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.926 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_15118_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_15118_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #2
File | emd_15118_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
-Entire : 13pf microtubule from recombinant human tubulin lacking the C-ter...
Entire | Name: 13pf microtubule from recombinant human tubulin lacking the C-terminal tail |
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Components |
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-Supramolecule #1: 13pf microtubule from recombinant human tubulin lacking the C-ter...
Supramolecule | Name: 13pf microtubule from recombinant human tubulin lacking the C-terminal tail type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Homo sapiens (human) |
-Macromolecule #1: Tubulin alpha-1B
Macromolecule | Name: Tubulin alpha-1B / type: protein_or_peptide / ID: 1 Details: aa range 1-439; internal 6xHis tag in 40-loop; C-terminal TTP-tag Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIHHHHHHGG GDDSFNTFFS ETGAGKHVPR AVFVDLEPTV IDEVRTGTYR QLFHPEQLIT GKEDAANNYA RGHYTIGKEI IDLVLDRIRK LADQCTGLQG FLVFHSFGGG TGSGFTSLLM ERLSVDYGKK ...String: MRECISIHVG QAGVQIGNAC WELYCLEHGI QPDGQMPSDK TIHHHHHHGG GDDSFNTFFS ETGAGKHVPR AVFVDLEPTV IDEVRTGTYR QLFHPEQLIT GKEDAANNYA RGHYTIGKEI IDLVLDRIRK LADQCTGLQG FLVFHSFGGG TGSGFTSLLM ERLSVDYGKK SKLEFSIYPA PQVSTAVVEP YNSILTTHTT LEHSDCAFMV DNEAIYDICR RNLDIERPTY TNLNRLISQI VSSITASLRF DGALNVDLTE FQTNLVPYPR IHFPLATYAP VISAEKAYHE QLSVAEITNA CFEPANQMVK CDPRHGKYMA CCLLYRGDVV PKDVNAAIAT IKTKRSIQFV DWCPTGFKVG INYQPPTVVP GGDLAKVQRA VCMLSNTTAI AEAWARLDHK FDLMYAKRAF VHWYVGEGME EGEFSEARED MAALEKDYEE VGVDSCLAYT GG |
-Macromolecule #2: Tubulin beta-3
Macromolecule | Name: Tubulin beta-3 / type: protein_or_peptide / ID: 2 / Details: C-terminal FLAG-tag / Enantiomer: LEVO |
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Source (natural) | Organism: Homo sapiens (human) |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: MREIVHIQAG QCGNQIGAKF WEVISDEHGI DPSGNYVGDS DLQLERISVY YNEASSHKYV PRAILVDLEP GTMDSVRSGA FGHLFRPDNF IFGQSGAGNN WAKGHYTEGA ELVDSVLDVV RKECENCDCL QGFQLTHSLG GGTGSGMGTL LISKVREEYP DRIMNTFSVV ...String: MREIVHIQAG QCGNQIGAKF WEVISDEHGI DPSGNYVGDS DLQLERISVY YNEASSHKYV PRAILVDLEP GTMDSVRSGA FGHLFRPDNF IFGQSGAGNN WAKGHYTEGA ELVDSVLDVV RKECENCDCL QGFQLTHSLG GGTGSGMGTL LISKVREEYP DRIMNTFSVV PSPKVSDTVV EPYNATLSIH QLVENTDETY CIDNEALYDI CFRTLKLATP TYGDLNHLVS ATMSGVTTSL RFPGQLNADL RKLAVNMVPF PRLHFFMPGF APLTARGSQQ YRALTVPELT QQMFDAKNMM AACDPRHGRY LTVATVFRGR MSMKEVDEQM LAIQSKNSSY FVEWIPNNVK VAVCDIPPRG LKMSSTFIGN STAIQELFKR ISEQFTAMFR RKAFLHWYTG EGMDEMEFTE AESNMNDLVS EYQQYQDATA EEEGEMYEDD EEESEAQGPK ENLYFQSSGG DYKDDDDK |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | helical reconstruction |
Aggregation state | helical array |
-Sample preparation
Buffer | pH: 6.8 |
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Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 50.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.6 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |