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- PDB-6rzb: Cryo-EM structure of mouse cytoplasmic dynein-1 microtubule bindi... -

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Entry
Database: PDB / ID: 6rzb
TitleCryo-EM structure of mouse cytoplasmic dynein-1 microtubule binding domain bound to microtubules
Components
  • MKIAA0325 protein
  • Tubulin alpha-1B chain
  • Tubulin beta chain
KeywordsMOTOR PROTEIN / filament / complex
Function / homology
Function and homology information


COPI-dependent Golgi-to-ER retrograde traffic / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Separation of Sister Chromatids / Resolution of Sister Chromatid Cohesion / HSP90 chaperone cycle for steroid hormone receptors (SHR) / Recruitment of NuMA to mitotic centrosomes / Hedgehog 'off' state / Cilium Assembly / Intraflagellar transport / RHO GTPases activate IQGAPs ...COPI-dependent Golgi-to-ER retrograde traffic / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Separation of Sister Chromatids / Resolution of Sister Chromatid Cohesion / HSP90 chaperone cycle for steroid hormone receptors (SHR) / Recruitment of NuMA to mitotic centrosomes / Hedgehog 'off' state / Cilium Assembly / Intraflagellar transport / RHO GTPases activate IQGAPs / COPI-mediated anterograde transport / Kinesins / Carboxyterminal post-translational modifications of tubulin / The role of GTSE1 in G2/M progression after G2 checkpoint / Mitotic Prometaphase / COPI-independent Golgi-to-ER retrograde traffic / RHO GTPases Activate Formins / dynein complex / microtubule motor activity / microtubule-based movement / microtubule-based process / structural constituent of cytoskeleton / microtubule cytoskeleton organization / neuron migration / mitotic cell cycle / microtubule / GTPase activity / GTP binding / cytoplasm
Dynein heavy chain AAA lid domain superfamily / Dynein heavy chain C-terminal domain / Dynein heavy chain, C-terminal domain / Dynein heavy chain AAA lid domain / Tubulin/FtsZ family, GTPase domain / Dynein heavy chain region D6 P-loop domain / Tubulin C-terminal domain / Microtubule-binding stalk of dynein motor / P-loop containing dynein motor region D4 / ATP-binding dynein motor region ...Dynein heavy chain AAA lid domain superfamily / Dynein heavy chain C-terminal domain / Dynein heavy chain, C-terminal domain / Dynein heavy chain AAA lid domain / Tubulin/FtsZ family, GTPase domain / Dynein heavy chain region D6 P-loop domain / Tubulin C-terminal domain / Microtubule-binding stalk of dynein motor / P-loop containing dynein motor region D4 / ATP-binding dynein motor region / Dynein heavy chain AAA lid domain / Tubulin subunits alpha, beta, and gamma signature. / Tubulin/FtsZ, GTPase domain superfamily / Tubulin-beta mRNA autoregulation signal. / Tubulin/FtsZ, C-terminal domain superfamily / Dynein heavy chain, ATP-binding dynein motor region / Tubulin/FtsZ, GTPase domain / Tubulin / P-loop containing nucleoside triphosphate hydrolase / Beta tubulin / Alpha tubulin / AAA+ ATPase domain / Dynein heavy chain region D6 P-loop domain / Tubulin/FtsZ, C-terminal / 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
Tubulin beta chain / Tubulin alpha-1B chain / MKIAA0325 protein
Biological speciesMus musculus (house mouse)
Sus scrofa (pig)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.1 Å
AuthorsLacey, S.E. / He, S. / Scheres, S.H.W. / Carter, A.P.
Funding supportUnited Kingdom , 2件
OrganizationGrant numberCountry
Medical Research Council (United Kingdom)MC_UP_A025_1011United Kingdom
Wellcome TrustWT210711United Kingdom
CitationJournal: Elife / Year: 2019
Title: Cryo-EM of dynein microtubule-binding domains shows how an axonemal dynein distorts the microtubule.
Authors: Samuel E Lacey / Shaoda He / Sjors Hw Scheres / Andrew P Carter /
Abstract: Dyneins are motor proteins responsible for transport in the cytoplasm and the beating of axonemes in cilia and flagella. They bind and release microtubules via a compact microtubule-binding domain ...Dyneins are motor proteins responsible for transport in the cytoplasm and the beating of axonemes in cilia and flagella. They bind and release microtubules via a compact microtubule-binding domain (MTBD) at the end of a coiled-coil stalk. We address how cytoplasmic and axonemal dynein MTBDs bind microtubules at near atomic resolution. We decorated microtubules with MTBDs of cytoplasmic dynein-1 and axonemal dynein DNAH7 and determined their cryo-EM structures using helical Relion. The majority of the MTBD is rigid upon binding, with the transition to the high affinity state controlled by the movement of a single helix at the MTBD interface. DNAH7 contains an 18-residue insertion, found in many axonemal dyneins, that contacts the adjacent protofilament. Unexpectedly we observe that DNAH7, but not dynein-1, induces large distortions in the microtubule cross-sectional curvature. This raises the possibility that dynein coordination in axonemes is mediated via conformational changes in the microtubule.
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Jun 13, 2019 / Release: Jul 10, 2019
RevisionDateData content typeGroupCategoryItemProviderType
1.0Jul 10, 2019Structure modelrepositoryInitial release
1.1Jul 17, 2019Structure modelData collection / Database referencescitation / citation_author_citation.journal_volume / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name

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Structure viewerMolecule:
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Assembly

Deposited unit
A: Tubulin alpha-1B chain
B: Tubulin beta chain
C: MKIAA0325 protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)115,2437
Polymers113,3983
Non-polymers1,8454
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area8970 Å2
ΔGint-47 kcal/mol
Surface area34880 Å2
MethodPISA

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Components

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Protein/peptide , 3 types, 3 molecules ABC

#1: Protein/peptide Tubulin alpha-1B chain / Alpha-tubulin ubiquitous / Tubulin K-alpha-1 / Tubulin alpha-ubiquitous chain


Mass: 48679.051 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / Organ: Brain / References: UniProt: Q2XVP4
#2: Protein/peptide Tubulin beta chain / Beta-tubulin


Mass: 47825.859 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Sus scrofa (pig) / References: UniProt: P02554
#3: Protein/peptide MKIAA0325 protein


Mass: 16893.574 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Dync1h1, Dnchc1, mKIAA0325 / Production host: Escherichia coli (E. coli) / References: UniProt: Q80U36

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Non-polymers , 4 types, 4 molecules

#4: Chemical ChemComp-GTP / GUANOSINE-5'-TRIPHOSPHATE


Mass: 523.180 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N5O14P3 / Guanosine triphosphate / Comment: GTP (energy-carrying molecule) *YM
#5: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg / Magnesium
#6: Chemical ChemComp-GDP / GUANOSINE-5'-DIPHOSPHATE


Mass: 443.201 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H15N5O11P2 / Guanosine diphosphate / Comment: GDP (energy-carrying molecule) *YM
#7: Chemical ChemComp-TA1 / TAXOL


Mass: 853.906 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C47H51NO14 / Paclitaxel

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Details

Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: HELICAL ARRAY / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component

Type: COMPLEX

IDNameEntity IDParent-IDSource
1Mouse cytoplasmic dynein-1 microtubule binding domain decorating microtubules1, 2, 30MULTIPLE SOURCES
2Tubulin1, 21NATURAL
3dynein-1 microtubule binding31NATURAL
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Sus scrofa (pig)9823
23Mus musculus (house mouse)10090
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 60 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

SoftwareName: REFMAC / Version: 5.8.0238 / Classification: refinement
EM software
IDNameVersionCategory
10RELION3final Euler assignment
12RELION33D reconstruction
13REFMAC5model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 59100 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT
RefinementResolution: 5→187.2 Å / Cor.coef. Fo:Fc: 0.532 / SU B: 171.433 / SU ML: 1.972 / ESU R: 1.061
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
RfactorNum. reflection% reflection
Rwork0.66163 --
Obs0.66163 109888 100 %
Solvent computationSolvent model: PARAMETERS FOR MASK CACLULATION
Displacement parametersBiso mean: 162.659 Å2
Baniso -1Baniso -2Baniso -3
1-5.51 Å21.22 Å2-0.51 Å2
2---1.29 Å2-1.62 Å2
3----4.23 Å2
Refinement stepCycle: 1 / Resolution: 5→187.2 Å / Total: 8001
Refine LS restraints

Refinement-ID: ELECTRON MICROSCOPY

TypeDev idealDev ideal targetNumber
r_bond_refined_d0.0060.0128178
r_bond_other_d
r_angle_refined_deg1.5121.64511109
r_angle_other_deg
r_dihedral_angle_1_deg7.1215997
r_dihedral_angle_2_deg32.02122.627434
r_dihedral_angle_3_deg18.471151370
r_dihedral_angle_4_deg16.6031550
r_chiral_restr0.1070.21073
r_gen_planes_refined0.0060.026291
r_gen_planes_other
r_nbd_refined
r_nbd_other
r_nbtor_refined
r_nbtor_other
r_xyhbond_nbd_refined
r_xyhbond_nbd_other
r_metal_ion_refined
r_metal_ion_other
r_symmetry_vdw_refined
r_symmetry_vdw_other
r_symmetry_hbond_refined
r_symmetry_hbond_other
r_symmetry_metal_ion_refined
r_symmetry_metal_ion_other
r_mcbond_it15.7314.4084000
r_mcbond_other
r_mcangle_it26.62221.564993
r_mcangle_other
r_scbond_it24.48117.3994178
r_scbond_other
r_scangle_it
r_scangle_other
r_long_range_B_refined58.60635231
r_long_range_B_other
r_rigid_bond_restr
r_sphericity_free
r_sphericity_bonded
LS refinement shellResolution: 5.001→5.131 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0 0 -
Rwork0.828 7966 -
Obs--100 %

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