|Entry||Database: PDB / ID: 3j8y|
|Title||High-resolution structure of ATP analog-bound kinesin on microtubules|
|Keywords||MOTOR PROTEIN/STRUCTURAL PROTEIN / molecular motors / kinesin / myosin / microtubules / cytoskeletal motors / MOTOR PROTEIN-STRUCTURAL PROTEIN complex|
|Function / homology||Kinesin motor domain / Kinesin-like protein / Resolution of Sister Chromatid Cohesion / HSP90 chaperone cycle for steroid hormone receptors (SHR) / Tubulin / Regulation of PLK1 Activity at G2/M Transition / Alpha tubulin / Beta tubulin / Tubulin/FtsZ, GTPase domain / Tubulin/FtsZ, C-terminal ...Kinesin motor domain / Kinesin-like protein / Resolution of Sister Chromatid Cohesion / HSP90 chaperone cycle for steroid hormone receptors (SHR) / Tubulin / Regulation of PLK1 Activity at G2/M Transition / Alpha tubulin / Beta tubulin / Tubulin/FtsZ, GTPase domain / Tubulin/FtsZ, C-terminal / Beta tubulin, autoregulation binding site / Tubulin, conserved site / Tubulin/FtsZ, 2-layer sandwich domain / Kinesin motor domain, conserved site / Tubulin, C-terminal / P-loop containing nucleoside triphosphate hydrolase / Tubulin/FtsZ, GTPase domain superfamily / Recruitment of mitotic centrosome proteins and complexes / Kinesin motor domain profile. / Microtubule-dependent trafficking of connexons from Golgi to the plasma membrane / Kinesins / COPI-dependent Golgi-to-ER retrograde traffic / RHO GTPases activate KTN1 / Insulin processing / MHC class II antigen presentation / Kinesin motor domain signature. / Kinesin motor domain superfamily / Tubulin-beta mRNA autoregulation signal. / Tubulin subunits alpha, beta, and gamma signature. / Tubulin C-terminal domain / Kinesin motor domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ, C-terminal domain superfamily / Loss of Nlp from mitotic centrosomes / Separation of Sister Chromatids / Loss of proteins required for interphase microtubule organization from the centrosome / Recycling pathway of L1 / Kinesins / Carboxyterminal post-translational modifications of tubulin / AURKA Activation by TPX2 / The role of GTSE1 in G2/M progression after G2 checkpoint / Mitotic Prometaphase / COPI-independent Golgi-to-ER retrograde traffic / COPI-dependent Golgi-to-ER retrograde traffic / COPI-mediated anterograde transport / RHO GTPases Activate Formins / RHO GTPases activate IQGAPs / Intraflagellar transport / Anchoring of the basal body to the plasma membrane / Cilium Assembly / Hedgehog 'off' state / Recruitment of NuMA to mitotic centrosomes / positive regulation of voltage-gated sodium channel activity / cytoplasm organization / plus-end-directed vesicle transport along microtubule / anterograde neuronal dense core vesicle transport / retrograde neuronal dense core vesicle transport / anterograde axonal protein transport / vesicle transport along microtubule / positive regulation of potassium ion transport / positive regulation of vesicle fusion / microtubule lateral binding / positive regulation of intracellular protein transport / stress granule disassembly / JUN kinase binding / regulation of synapse organization / ciliary rootlet / centrosome localization / spindle assembly / kinesin complex / MHC class I protein binding / microtubule motor activity / nuclear envelope lumen / positive regulation of insulin secretion involved in cellular response to glucose stimulus / axon cytoplasm / microtubule-based movement / microtubule-based process / phagocytic vesicle / regulation of membrane potential / axonal growth cone / cytoplasmic ribonucleoprotein granule / cellular response to interferon-gamma / structural constituent of cytoskeleton / microtubule cytoskeleton organization / hippocampus development / neuron migration / positive regulation of synaptic transmission, GABAergic / microtubule organizing center / positive regulation of protein localization to plasma membrane / cell body / mitotic cell cycle / microtubule / microtubule binding / vesicle / ATPase activity / cadherin binding / GTPase activity / GTP binding / ubiquitin protein ligase binding / perinuclear region of cytoplasm|
Function and homology information
|Specimen source||Homo sapiens (human)|
Sus scrofa (pig)
|Method||ELECTRON MICROSCOPY / helical reconstruction / cryo EM / 5 Å resolution|
|Authors||Shang, Z. / Zhou, K. / Xu, C. / Csencsits, R. / Cochran, J.C. / Sindelar, C.V.|
|Citation||Journal: Elife / Year: 2014|
Title: High-resolution structures of kinesin on microtubules provide a basis for nucleotide-gated force-generation.
Authors: Zhiguo Shang / Kaifeng Zhou / Chen Xu / Roseann Csencsits / Jared C Cochran / Charles V Sindelar
SummaryFull reportAbout validation report
|Date||Deposition: Nov 20, 2014 / Release: Dec 10, 2014|
|Structure viewer||Molecule: |
Downloads & links
K: Kinesin-1 heavy chain
A: Tubulin alpha-1B chain
B: Tubulin beta-2B chain
|Helical symmetry||Circular symmetry: 1 / Dyad axis: no / N subunits divisor: 1 / Number of operations: 1 / Rise per n subunits: 8.5215 Å / Rotation per n subunits: -25.77 deg.|
|Details||The reconstructed 14-protofilament microtubule is pseudo-symmetric, containing a seam with 3 starts per tubulin monomer, or 1.5 starts per tubulin dimer.|
-Protein/peptide , 3 types, 3 molecules K
|#1: Protein/peptide|| |
Mass: 39238.145 Da / Num. of mol.: 1
Fragment: Truncated catalytic head domain (monomeric, UNP residues 1-349)
Source: (gene. exp.) Homo sapiens (human) / Gene: KIF5B, KNS, KNS1 / Production host: Escherichia coli (E. coli) / References: UniProt: P33176
|#2: Protein/peptide|| |
Mass: 50204.445 Da / Num. of mol.: 1 / Source: (natural) Sus scrofa (pig) / References: UniProt: Q2XVP4
|#3: Protein/peptide|| |
Mass: 49983.824 Da / Num. of mol.: 1 / Source: (natural) Sus scrofa (pig) / References: UniProt: F2Z5B2
-Non-polymers , 4 types, 4 molecules
|#4: Chemical|| ChemComp-ATP / |
|#5: Chemical|| ChemComp-MG / |
|#6: Chemical|| ChemComp-GTP / |
|#7: Chemical|| ChemComp-GDP / |
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: FILAMENT / Reconstruction method: helical reconstruction|
|Molecular weight||Value: 0.135 MDa / Experimental value: NO|
|Buffer solution||Name: 25 mM PIPES, 25 mM NaCl, 2 mM MgCl2, 1 mM EGTA / Details: 25 mM PIPES, 25 mM NaCl, 2 mM MgCl2, 1 mM EGTA / pH: 6.8|
|Specimen||Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Details: 300 mesh copper grid with homemade holey carbon|
|Vitrification||Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE|
Details: No glow discharge was applied. After sample application to grid, liquid was mostly 'wicked' away by edgewise application of filter paper. Subsequently, blotting and plunge freezing were performed with ~0.5 second delay after blotting but prior to plunging into liquid ethane.
Method: No glow discharge was applied. After sample application to grid, liquid was mostly 'wicked' away by edgewise application of filter paper. Subsequently, blotting and plunge freezing were performed with ~0.5 second delay after blotting but prior to plunging.
-Electron microscopy imaging
|Microscopy||Microscope model: FEI TITAN / Date: Jun 2, 2013|
Details: 4K x 4K counting mode was used. 24 frames total were collected.
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELDBright-field microscopy / Calibrated magnification: 23859 / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Cs: 2 mm / Camera length: 0 mm|
|Specimen holder||Specimen holder model: GATAN LIQUID NITROGEN|
|Image recording||Electron dose: 15 e/Å2 / Film or detector model: GATAN ULTRASCAN 4000 (4k x 4k)|
|Image scans||Number digital images: 51|
|Radiation||Diffraction protocol: SINGLE WAVELENGTH / Monochromatic or laue m l: M / Scattering type: x-ray|
|Radiation wavelength||Relative weight: 1|
|CTF correction||Details: done within FREALIGN|
|Helical symmerty||Angular rotation/subunit: 25.77 deg. / Axial rise/subunit: 8.5215 Å / Axial symmetry: C1|
Details: The reconstructed 14-protofilament microtubule is pseudo-symmetric, containing a seam with 3 starts per tubulin monomer, or 1.5 starts per tubulin dimer.
|3D reconstruction||Method: Single particleSingle particle analysis / Resolution: 5 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 49961 / Nominal pixel size: 2.097 / Actual pixel size: 2.097|
Details: Initial alignment was done using customized SPIDER scripts. Reconstruction and subsequent refinement were done by FREALIGN.
Symmetry type: HELICAL
|Atomic model building||Details: REFINEMENT PROTOCOL--flexible DETAILS--MDFF was performed using explicit solvation, after placing active-site water coordinates identified in high-resolution crystal structures of kinesins ATP-like state. Side chains were excluded from the MDFF target potential. Following several equilibration steps, the relative strength of the EM map potential (GSCALE term) was slowly increased from 0 to 1 over the course of 10 nanoseconds. The t = 1.4 ns time point was selected to represent the final fitted model, based on the approximate convergence of the RMSD from the starting structure.|
Ref protocol: FLEXIBLE FIT / Ref space: REAL
Target criteria: RMSD from the starting structure was monitored for convergence.
|Atomic model building|
|Number of atoms included #LAST||Protein: 9343 / Nucleic acid: 0 / Ligand: 92 / Solvent: 0 / Total: 9435|
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