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- PDB-3j6p: Pseudo-atomic model of dynein microtubule binding domain-tubulin ... -

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Basic information

Entry
Database: PDB / ID: 3j6p
TitlePseudo-atomic model of dynein microtubule binding domain-tubulin complex based on a cryoEM map
Components
  • Dynein heavy chain, cytoplasmic
  • Tubulin alpha-1A chain
  • Tubulin beta chain
KeywordsMOTOR PROTEIN/STRUCTURAL PROTEIN / Motor Protein-Cytoskeleton Complex / MOTOR PROTEIN-STRUCTURAL PROTEIN complex
Function / homology
Function and homology information


phagolysosome membrane / early phagosome membrane / CTPase activity / phagosome maturation / minus-end-directed vesicle transport along microtubule / establishment of spindle localization / ATP-dependent microtubule motor activity, minus-end-directed / dynein intermediate chain binding / dynein light intermediate chain binding / cytoplasmic dynein complex ...phagolysosome membrane / early phagosome membrane / CTPase activity / phagosome maturation / minus-end-directed vesicle transport along microtubule / establishment of spindle localization / ATP-dependent microtubule motor activity, minus-end-directed / dynein intermediate chain binding / dynein light intermediate chain binding / cytoplasmic dynein complex / dynein complex / nuclear migration / cytoplasmic microtubule organization / microtubule-based movement / cytoplasmic microtubule / mitotic spindle assembly / microtubule-based process / tubulin binding / endocytic vesicle / structural constituent of cytoskeleton / neuron migration / microtubule cytoskeleton organization / cell cortex / mitotic cell cycle / microtubule binding / microtubule / ATPase activity / GTPase activity / centrosome / GTP binding / ATP binding / identical protein binding / cytoplasm
Tubulin C-terminal domain / P-loop containing nucleoside triphosphate hydrolase / Tubulin/FtsZ, GTPase domain / AAA+ ATPase domain / Dynein heavy chain region D6 P-loop domain / Tubulin/FtsZ, C-terminal / Dynein heavy chain, domain-1 / Dynein heavy chain, domain-2 / Beta tubulin, autoregulation binding site / Tubulin, conserved site ...Tubulin C-terminal domain / P-loop containing nucleoside triphosphate hydrolase / Tubulin/FtsZ, GTPase domain / AAA+ ATPase domain / Dynein heavy chain region D6 P-loop domain / Tubulin/FtsZ, C-terminal / Dynein heavy chain, domain-1 / Dynein heavy chain, domain-2 / Beta tubulin, autoregulation binding site / Tubulin, conserved site / Tubulin/FtsZ, 2-layer sandwich domain / Tubulin, C-terminal / Dynein heavy chain, AAA module D4 / Dynein heavy chain, coiled coil stalk / Dynein heavy chain, hydrolytic ATP-binding dynein motor region / Alpha tubulin / Dynein heavy chain, ATP-binding dynein motor region / Tubulin/FtsZ, GTPase domain superfamily / Tubulin/FtsZ, C-terminal domain superfamily / Dynein heavy chain, C-terminal domain / Dynein heavy chain, AAA 5 extension domain / Dynein heavy chain 3, AAA+ lid domain / Dynein heavy chain AAA lid domain / Dynein heavy chain AAA lid domain superfamily / Dynein heavy chain, domain 2, N-terminal / Dynein heavy chain, domain 2, C-terminal / Dynein heavy chain, AAA1 domain, small subdomain / Dynein heavy chain, C-terminal domain, barrel region / Tubulin/FtsZ family, GTPase domain / Beta tubulin / Tubulin
Tubulin alpha-1A chain / Tubulin beta chain / Dynein heavy chain, cytoplasmic
Biological speciesDictyostelium discoideum (eukaryote)
Sus scrofa (pig)
MethodELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 8.2 Å
AuthorsUchimura, S. / Fujii, T. / Takazaki, H. / Ayukawa, R. / Nishikawa, Y. / Minoura, I. / Hachikubo, Y. / Kurisu, G. / Sutoh, K. / Kon, T. ...Uchimura, S. / Fujii, T. / Takazaki, H. / Ayukawa, R. / Nishikawa, Y. / Minoura, I. / Hachikubo, Y. / Kurisu, G. / Sutoh, K. / Kon, T. / Namba, K. / Muto, E.
CitationJournal: J Cell Biol / Year: 2015
Title: A flipped ion pair at the dynein-microtubule interface is critical for dynein motility and ATPase activation.
Authors: Seiichi Uchimura / Takashi Fujii / Hiroko Takazaki / Rie Ayukawa / Yosuke Nishikawa / Itsushi Minoura / You Hachikubo / Genji Kurisu / Kazuo Sutoh / Takahide Kon / Keiichi Namba / Etsuko Muto /
Abstract: Dynein is a motor protein that moves on microtubules (MTs) using the energy of adenosine triphosphate (ATP) hydrolysis. To understand its motility mechanism, it is crucial to know how the signal of ...Dynein is a motor protein that moves on microtubules (MTs) using the energy of adenosine triphosphate (ATP) hydrolysis. To understand its motility mechanism, it is crucial to know how the signal of MT binding is transmitted to the ATPase domain to enhance ATP hydrolysis. However, the molecular basis of signal transmission at the dynein-MT interface remains unclear. Scanning mutagenesis of tubulin identified two residues in α-tubulin, R403 and E416, that are critical for ATPase activation and directional movement of dynein. Electron cryomicroscopy and biochemical analyses revealed that these residues form salt bridges with the residues in the dynein MT-binding domain (MTBD) that work in concert to induce registry change in the stalk coiled coil and activate the ATPase. The R403-E3390 salt bridge functions as a switch for this mechanism because of its reversed charge relative to other residues at the interface. This study unveils the structural basis for coupling between MT binding and ATPase activation and implicates the MTBD in the control of directional movement.
Validation Report
SummaryFull reportAbout validation report
History
DepositionMar 20, 2014Deposition site: RCSB / Processing site: PDBJ
Revision 1.0Dec 31, 2014Provider: repository / Type: Initial release
Revision 1.1Apr 8, 2015Group: Database references

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Structure visualization

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Structure viewerMolecule:
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Assembly

Deposited unit
D: Dynein heavy chain, cytoplasmic
A: Tubulin alpha-1A chain
B: Tubulin beta chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)114,1877
Polymers112,3433
Non-polymers1,8454
Water0
1


TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Protein , 3 types, 3 molecules DAB

#1: Protein Dynein heavy chain, cytoplasmic / Dynein heavy chain / cytosolic / DYHC


Mass: 12327.559 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Dictyostelium discoideum (eukaryote) / Gene: DDB_G0276355, dhcA / Production host: Escherichia coli (E. coli) / References: UniProt: P34036
#2: Protein Tubulin alpha-1A chain / Alpha-tubulin 1 / Tubulin alpha-1 chain


Mass: 50107.238 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / References: UniProt: P02550
#3: Protein Tubulin beta chain / Beta-tubulin


Mass: 49907.770 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / References: UniProt: P02554

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Non-polymers , 4 types, 4 molecules

#4: Chemical ChemComp-MG / MAGNESIUM ION / Magnesium


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg
#5: Chemical ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE / Guanosine triphosphate


Mass: 523.180 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O14P3 / Comment: GTP, energy-carrying molecule*YM
#6: Chemical ChemComp-GDP / GUANOSINE-5'-DIPHOSPHATE / Guanosine diphosphate


Type: RNA linking / Mass: 443.201 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H15N5O11P2 / Comment: GDP, energy-carrying molecule*YM
#7: Chemical ChemComp-TA1 / TAXOL / Paclitaxel


Mass: 853.906 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C47H51NO14 / Comment: medication, chemotherapy*YM

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: FILAMENT / 3D reconstruction method: helical reconstruction

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Sample preparation

ComponentName: Dictyostelium dynein microtubule binding domain bound to a 15-protofilament microtubule
Type: COMPLEX
Buffer solutionpH: 7
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK I / Cryogen name: ETHANE / Humidity: 100 % / Method: Blot for 3.5 seconds before plunging

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Electron microscopy imaging

MicroscopyModel: JEOL 3200FSC / Date: Nov 5, 2012
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 60000 X / Calibrated magnification: 109489 X / Nominal defocus max: 1500 nm / Nominal defocus min: 1000 nm / Cs: 1.6 mm
Specimen holderTemperature: 50 K
Image recordingElectron dose: 20 e/Å2 / Film or detector model: TVIPS TEMCAM-F415 (4k x 4k)
Radiation wavelengthRelative weight: 1

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Processing

EM software
IDNameCategory
1DireXmodel fitting
2FlexEMmodel fitting
3SPIDER3D reconstruction
CTF correctionDetails: CTFFIND3 Each particle
3D reconstructionMethod: Projection Matching / Resolution: 8.2 Å / Actual pixel size: 1.37 Å / Symmetry type: HELICAL
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL / Target criteria: Cross-correlation coefficient
Details: METHOD--Flexible fitting REFINEMENT PROTOCOL--Flexible fitting
Atomic model building
IDPDB-ID3D fitting-ID
13VKH1
21JFF1
Refinement stepCycle: LAST
ProteinNucleic acidLigandSolventTotal
Num. atoms7440 0 123 0 7563

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