Near-atomic cryo-EM structure of yeast kinesin-5-microtubule complex reveals a distinct binding footprint

Kinesin-5s are essential members of the superfamily of microtubule-dependent motors that undertake conserved roles in cell division. We investigated coevolution of the motor-microtubule interface using cryo-electron microscopy to determine the near-atomic structure of the motor domain of Cut7, the fission yeast kinesin-5, bound to fission yeast microtubules. AMPPNP-bound Cut7 adopts a kinesin-conserved ATP-like conformation, with a closed nucleotide binding pocket and docked neck linker that supports cover neck bundle formation. Compared to mammalian tubulin microtubules, Cut7’s footprint on S. pombe microtubule surface is subtly different because of their different architecture. However, the core motor-microtubule interaction that stimulates motor ATPase is tightly conserved, reflected in similar Cut7 ATPase activities on each microtubule type. The S. pombe microtubules were bound by the drug epothilone, which is visible in the taxane binding pocket. Stabilization of S. pombe microtubules is mediated by drug binding at this conserved site despite their noncanonical architecture and mechanochemistry. Highlights S. pombe Cut7 has a distinct binding footprint on S. pombe microtubules The core interface driving microtubule activation of motor ATPase is conserved The neck linker is docked in AMPPNP-bound Cut7 and the cover neck bundle is formed Epothilone binds at the taxane binding site to stabilize S. pombe microtubules eTOC text To investigate coevolution of the motor-microtubule interface, we used cryo-electron microscopy to determine the near-atomic structure of the motor domain of Cut7, the fission yeast kinesin-5, bound to microtubules polymerized from natively purified fission yeast tubulin and stabilised by the drug epothilone.

• Publication year

  2018

• Venue

  bioRxiv

• Publication date

  2018-04-16

• Fields of study

  Biology, Chemistry

• Identifiers
  DOI 10.1101/302455
• External record

  Open on Semantic Scholar

• Source metadata

  Semantic Scholar

• No claims are published for this paper.

• No concepts are published for this paper.

• Nucleotide– and Mal3-dependent changes in fission yeast microtubules suggest a structural plasticity view of dynamics

  2017influential reference
• The yeast kinesin-5 Cin8 interacts with the microtubule in a noncanonical manner

  2017cited by this paper
• Insights into the Distinct Mechanisms of Action of Taxane and Non-Taxane Microtubule Stabilizers from Cryo-EM Structures.

  2017cited by this paper
• Structural differences between yeast and mammalian microtubules revealed by cryo-EM

  2017cited by this paper
• Schizosaccharomyces pombe kinesin-5 switches direction using a steric blocking mechanism

  2016influential reference
• Scipion: A software framework toward integration, reproducibility and validation in 3D electron microscopy.

  2016cited by this paper
• High-resolution structures of kinesin on microtubules provide a basis for nucleotide-gated force-generation

  2014cited by this paper
• Conserved mechanisms of microtubule-stimulated ADP release, ATP binding, and force generation in transport kinesins

  2014cited by this paper
• Regulation of microtubule motors by tubulin isotypes and posttranslational modifications

  2014cited by this paper
• High-resolution microtubule structures reveal the structural transitions in αβ-tubulin upon GTP hydrolysis.

  2014cited by this paper
• Bidirectional motility of the fission yeast kinesin-5, Cut7.

  2014cited by this paper
• One number does not fit all: mapping local variations in resolution in cryo-EM reconstructions.

  2013cited by this paper
• Molecular Mechanism of Action of Microtubule-Stabilizing Anticancer Agents

  2013influential reference
• New insights into the mechanism of force generation by kinesin-5 molecular motors.

  2013influential reference
• Overexpression, purification, and functional analysis of recombinant human tubulin dimer

  2013cited by this paper
• Kinesin-5 Kip1 is a bi-directional motor that stabilizes microtubules and tracks their plus-ends in vivo

  2013cited by this paper
• Structure of a kinesin–tubulin complex and implications for kinesin motility

  2013cited by this paper
• The Structural Basis of Force Generation by the Mitotic Motor Kinesin-5*

  2012cited by this paper
• RELION: Implementation of a Bayesian approach to cryo-EM structure determination

  2012cited by this paper
• One-step purification of assembly-competent tubulin from diverse eukaryotic sources

  2012cited by this paper
• Loop L 5 Acts as a Conformational Latch in the Mitotic Kinesin

  2011cited by this paper
• Purification of tubulin from the fission yeast Schizosaccharomyces pombe.

  2011cited by this paper
• Directionality of individual kinesin‐5 Cin8 motors is modulated by loop 8, ionic strength and microtubule geometry

  2011cited by this paper
• Directional Switching of the Kinesin Cin8 Through Motor Coupling

  2011cited by this paper
• Loop 5-directed Compounds Inhibit Chimeric Kinesin-5 Motors

  2010cited by this paper
• Loop L5 Acts as a Conformational Latch in the Mitotic Kinesin Eg5*

  2010cited by this paper
• An atomic-level mechanism for activation of the kinesin molecular motors

  2010cited by this paper
• ATP Hydrolysis in Eg5 Kinesin Involves a Catalytic Two-water Mechanism*♦

  2009influential reference
• 9-Angström structure of a microtubule-bound mitotic motor.

  2009cited by this paper
• Kinesin's cover-neck bundle folds forward to generate force

  2008cited by this paper
• The beginning of kinesin's force-generating cycle visualized at 9-Å resolution

  2007cited by this paper
• Identification of a strong binding site for kinesin on the microtubule using mutant analysis of tubulin

  2006cited by this paper
• The bipolar mitotic kinesin Eg5 moves on both microtubules that it crosslinks

  2005cited by this paper
• UCSF Chimera—A visualization system for exploratory research and analysis

  2004cited by this paper
• research papers Acta Crystallographica Section D Biological

  2003cited by this paper
• Epothilone and paclitaxel: unexpected differences in promoting the assembly and stabilization of yeast microtubules.

  2002cited by this paper
• T-Coffee: A novel method for fast and accurate multiple sequence alignment.

  2000cited by this paper
• Correspondence e-mail:

  2000cited by this paper
• Asparagine and glutamine: using hydrogen atom contacts in the choice of side-chain amide orientation.

  1999cited by this paper
• EMAN: semiautomated software for high-resolution single-particle reconstructions.

  1999cited by this paper
• A structural change in the kinesin motor protein that drives motility

  1999cited by this paper
• Computer visualization of three-dimensional image data using IMOD.

  1996cited by this paper
• Structural changes accompanying GTP hydrolysis in microtubules: information from a slowly hydrolyzable analogue guanylyl-(alpha,beta)- methylene-diphosphonate

  1995cited by this paper
• Tubulin GTP hydrolysis influences the structure, mechanical properties, and kinesin-driven transport of microtubules.

  1994cited by this paper
• Yeast proteins associated with microtubules in vitro and in vivo.

  1992cited by this paper
• Novel potential mitotic motor protein encoded by the fission yeast cut7+ gene

  1990cited by this paper
• Escherichia coli phage T4 topoisomerase.

  1983cited by this paper
• Electronic Reprint Biological Crystallography Phenix : a Comprehensive Python-based System for Macromolecular Structure Solution

  year unknowncited by this paper

• No citing papers are available for this paper.