Crystal structure of Zen4 in the apo state reveals a missing conformation of kinesin

Ruifang Guan,Keith J Mickolajczyk,Yongfang Zhao,Zhucheng Chen,Lei Zhang,Geng-Yuan Chen,Yujie Sun,William O Hancock,Qian Peter Su

doi:10.1038/ncomms14951

Abstract

Kinesins hydrolyse ATP to transport intracellular cargoes along microtubules. Kinesin neck linker (NL) functions as the central mechano-chemical coupling element by changing its conformation through the ATPase cycle. Here we report the crystal structure of kinesin-6 Zen4 in a nucleotide-free, apo state, with the NL initial segment (NIS) adopting a backward-docked conformation and the preceding α6 helix partially melted. Single-molecule fluorescence resonance energy transfer (smFRET) analyses indicate the NIS of kinesin-1 undergoes similar conformational changes under tension in the two-head bound (2HB) state, whereas it is largely disordered without tension. The backward-docked structure of NIS is essential for motility of the motor. Our findings reveal a key missing conformation of kinesins, which provides the structural basis of the stable 2HB state and offers a tension-based rationale for an optimal NL length to ensure processivity of the motor.

Highlights

Kinesins hydrolyse ATP to transport intracellular cargoes along microtubules
The ATPase cycle of kinesin consists of three principle states: ATP, ADP and apo states
Our structure of Zen[4] shows that the NL initial segment (NIS) adopts a backward-docked conformation in the apo state. Single-molecule fluorescence resonance energy transfer (smFRET) analysis suggests the NIS of kinesin-1 adopts a similar conformation under tension, instead of being disordered as generally believed before

Summary

Introduction

Kinesins hydrolyse ATP to transport intracellular cargoes along microtubules. Kinesin neck linker (NL) functions as the central mechano-chemical coupling element by changing its conformation through the ATPase cycle. Very few structures of kinesin in the apo state have been reported[20,21,22], as the motor domain by itself is unstable in the absence of nucleotide[23] It cannot be measured, the magnitude of intramolecular tension can be estimated by the worm-like chain (WLC) model, which has been shown to faithfully recapitulate the force-extension curves of unfolded polypeptide and DNA24,25. Considering the stable binding of kinesin heads in the 2HB state, these magnitudes are large relative to the B7 pN unbinding force or stall force, and seem improbable[27,28,29,30] One reason for this discrepancy is that remains unclear what conformation the NL and the preceding a6 helix adopt under tension in the apo state[27]

Methods

Results

Conclusion