The kinesin-5 chemomechanical cycle is dominated by a two-heads-bound state

Geng Yuan Chen, Keith J. Mickolajczyk, William O. Hancock

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22 Scopus citations


Single-molecule microscopy and stopped-flow kinetics assays were carried out to understand the microtubule polymerase activity of kinesin-5 (Eg5). Four lines of evidence argue that the motor primarily resides in a two-heads-bound (2HB) state. First, upon microtubule binding, dimeric Eg5 releases both bound ADPs. Second, microtubule dissociation in saturating ADP is 20-fold slower for the dimer than for the monomer. Third, ATPtriggered mant-ADP release is 5-fold faster than the stepping rate. Fourth, ATP binding is relatively fast when the motor is locked in a 2HB state. Shortening the neck-linker does not facilitate rear-head detachment, suggesting a minimal role for rearhead-gating. This 2HB state may enable Eg5 to stabilize incoming tubulin at the growing microtubule plus-end. The finding that slowly hydrolyzable ATP analogs trigger slower nucleotide release than ATP suggests that ATP hydrolysis in the bound head precedes stepping by the tethered head, leading to a mechanochemical cycle in which processivity is determined by the race between unbinding of the bound head and attachment of the tethered head.

Original languageEnglish (US)
Pages (from-to)20283-20294
Number of pages12
JournalJournal of Biological Chemistry
Issue number39
StatePublished - Sep 23 2016

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology


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