|Entry||Database: PDB / ID: 4ck7|
|Title||Pseudo-atomic model of microtubule-bound human kinesin-5 motor domain in presence of adp.alfx (NECK-LINKER IN ITS DISCONNECTED CONFORMATION, based on cryo-electron microscopy experiment|
|Keywords||MOTOR PROTEIN / KINESINS / MICROTUBULES / MITOSIS / MECHANOCHEMISTRY|
|Function / homology|
Function and homology information
Kinesins / plus-end-directed microtubule motor activity / positive regulation of axon guidance / regulation of mitotic centrosome separation / COPI-dependent Golgi-to-ER retrograde traffic / mitotic centrosome separation / microtubule motor activity / kinesin complex / spindle organization / mitotic spindle assembly ...Kinesins / plus-end-directed microtubule motor activity / positive regulation of axon guidance / regulation of mitotic centrosome separation / COPI-dependent Golgi-to-ER retrograde traffic / mitotic centrosome separation / microtubule motor activity / kinesin complex / spindle organization / mitotic spindle assembly / microtubule-based movement / microtubule-based process / MHC class II antigen presentation / mitotic spindle organization / mitotic spindle / microtubule cytoskeleton organization / structural constituent of cytoskeleton / microtubule cytoskeleton / spindle / spindle pole / nervous system development / mitotic cell cycle / microtubule binding / microtubule / protein heterodimerization activity / cell division / GTP binding / protein kinase binding / protein-containing complex / membrane / ATP binding / nucleus / cytosol / cytoplasm
Similarity search - Function
Kinesin-associated microtubule-binding / Kinesin-associated microtubule-binding domain / Kinesin motor domain signature. / Kinesin motor domain, conserved site / Kinesin motor, catalytic domain. ATPase. / Kinesin motor domain / Kinesin motor domain profile. / Kinesin motor domain / Tubulin-beta mRNA autoregulation signal. / Beta tubulin, autoregulation binding site ...Kinesin-associated microtubule-binding / Kinesin-associated microtubule-binding domain / Kinesin motor domain signature. / Kinesin motor domain, conserved site / Kinesin motor, catalytic domain. ATPase. / Kinesin motor domain / Kinesin motor domain profile. / Kinesin motor domain / Tubulin-beta mRNA autoregulation signal. / Beta tubulin, autoregulation binding site / Beta tubulin / Alpha tubulin / Kinesin motor domain superfamily / Tubulin C-terminal domain / Tubulin, C-terminal / Tubulin / Tubulin, conserved site / Tubulin subunits alpha, beta, and gamma signature. / Tubulin/FtsZ family, C-terminal domain / Tubulin/FtsZ-like, C-terminal domain / Tubulin/FtsZ, 2-layer sandwich domain / Tubulin/FtsZ, C-terminal / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ family, GTPase domain / Tubulin/FtsZ, GTPase domain / Tubulin/FtsZ, GTPase domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Kinesin-like protein KIF11 / Tubulin alpha-1D chain / ADENOSINE-5'-DIPHOSPHATE / ALUMINUM FLUORIDE / TAXOL / GUANOSINE-5'-TRIPHOSPHATE / GUANOSINE-5'-DIPHOSPHATE / Tubulin beta-2B chain
Similarity search - Component
|Biological species||HOMO SAPIENS (human)|
BOS TAURUS (cattle)
|Method||ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 9.2 Å|
|Model type details||CA ATOMS ONLY, CHAIN A, B, C|
|Authors||Goulet, A. / Major, J. / Jun, Y. / Gross, S. / Rosenfeld, S. / Moores, C.|
|Citation||Journal: Proc Natl Acad Sci U S A / Year: 2014|
Title: Comprehensive structural model of the mechanochemical cycle of a mitotic motor highlights molecular adaptations in the kinesin family.
Authors: Adeline Goulet / Jennifer Major / Yonggun Jun / Steven P Gross / Steven S Rosenfeld / Carolyn A Moores /
Abstract: Kinesins are responsible for a wide variety of microtubule-based, ATP-dependent functions. Their motor domain drives these activities, but the molecular adaptations that specify these diverse and ...Kinesins are responsible for a wide variety of microtubule-based, ATP-dependent functions. Their motor domain drives these activities, but the molecular adaptations that specify these diverse and essential cellular activities are poorly understood. It has been assumed that the first identified kinesin--the transport motor kinesin-1--is the mechanistic paradigm for the entire superfamily, but accumulating evidence suggests otherwise. To address the deficits in our understanding of the molecular basis of functional divergence within the kinesin superfamily, we studied kinesin-5s, which are essential mitotic motors whose inhibition blocks cell division. Using cryo-electron microscopy and determination of structure at subnanometer resolution, we have visualized conformations of microtubule-bound human kinesin-5 motor domain at successive steps in its ATPase cycle. After ATP hydrolysis, nucleotide-dependent conformational changes in the active site are allosterically propagated into rotations of the motor domain and uncurling of the drug-binding loop L5. In addition, the mechanical neck-linker element that is crucial for motor stepping undergoes discrete, ordered displacements. We also observed large reorientations of the motor N terminus that indicate its importance for kinesin-5 function through control of neck-linker conformation. A kinesin-5 mutant lacking this N terminus is enzymatically active, and ATP-dependent neck-linker movement and motility are defective, although not ablated. All these aspects of kinesin-5 mechanochemistry are distinct from kinesin-1. Our findings directly demonstrate the regulatory role of the kinesin-5 N terminus in collaboration with the motor's structured neck-linker and highlight the multiple adaptations within kinesin motor domains that tune their mechanochemistries according to distinct functional requirements.
|Structure viewer||Molecule: |
Downloads & links
A: TUBULIN ALPHA-1D CHAIN
B: TUBULIN BETA-2B CHAIN
C: KINESIN-LIKE PROTEIN KIF11
-Protein , 3 types, 3 molecules A
|#1: Protein|| |
Mass: 50236.352 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) BOS TAURUS (cattle) / Organ: BRAIN / References: UniProt: Q2HJ86
|#2: Protein|| |
Mass: 49907.770 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) BOS TAURUS (cattle) / Organ: BRAIN / References: UniProt: Q6B856
|#3: Protein|| |
Mass: 41673.105 Da / Num. of mol.: 1 / Fragment: MOTOR DOMAIN, RESIDUES 1-367 / Mutation: YES
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) HOMO SAPIENS (human) / Plasmid: PET21A / Production host: ESCHERICHIA COLI (E. coli) / Strain (production host): BL21 / References: UniProt: P52732
-Non-polymers , 6 types, 7 molecules
|#4: Chemical||#5: Chemical|| ChemComp-GTP / ||#6: Chemical|| ChemComp-GDP / ||#7: Chemical|| ChemComp-TA1 / ||#8: Chemical|| ChemComp-AF3 / ||#9: Chemical|| ChemComp-ADP / |
|Sequence details||CYS-LITE MUTANT CONTAINING|
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction|
|Component||Name: MICROTUBULE-BOUND HUMAN KINESIN-5 MOTOR DOMAIN / Type: COMPLEX|
|Buffer solution||Name: 80 MM PIPES 5 MM MGCL2 1 MM EGTA 1 MM TCEP / pH: 6.8 / Details: 80 MM PIPES 5 MM MGCL2 1 MM EGTA 1 MM TCEP|
|Specimen||Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Details: HOLEY CARBON|
|Vitrification||Instrument: FEI VITROBOT MARK I / Cryogen name: ETHANE / Details: LIQUID ETHANE|
-Electron microscopy imaging
Model: Tecnai F20 / Image courtesy: FEI Company
|Microscopy||Model: FEI TECNAI F20 / Date: Mar 10, 2011|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM|
|Electron lens||Mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 50000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 700 nm / Cs: 2 mm|
|Specimen holder||Temperature: 90 K|
|Image recording||Electron dose: 18 e/Å2 / Film or detector model: KODAK SO-163 FILM|
|Image scans||Num. digital images: 125|
|CTF correction||Details: FREALIGN|
|Symmetry||Point symmetry: C1 (asymmetric)|
|3D reconstruction||Resolution: 9.2 Å / Num. of particles: 10692 |
Details: THE N-TERMINAL SEQUENCE WAS MODELLED USING MODELLER. THE DISCONNECTED CONFORMATION OF THE NECK-LINKER WAS CALCULATED USING A CONJUGATE-GRADIENT ENERGY MINIMIZATION. SUBMISSION BASED ON ...Details: THE N-TERMINAL SEQUENCE WAS MODELLED USING MODELLER. THE DISCONNECTED CONFORMATION OF THE NECK-LINKER WAS CALCULATED USING A CONJUGATE-GRADIENT ENERGY MINIMIZATION. SUBMISSION BASED ON EXPERIMENTAL DATA FROM EMDB EMD-2533. (DEPOSITION ID: 12198).
Symmetry type: POINT
|Atomic model building||Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: Cross-correlation coefficient / Details: METHOD--FLEXIBLE REFINEMENT PROTOCOL--X-RAY|
|Atomic model building|
|Refinement||Highest resolution: 9.2 Å|
|Refinement step||Cycle: LAST / Highest resolution: 9.2 Å|
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