The beginning of kinesin's force-generating cycle visualized at 9-A resolution

Area of Science:

• Biochemistry
• Structural Biology
• Molecular Motors

Background:

• Kinesin is a crucial motor protein that transports cargo along microtubules.
• Understanding kinesin's nucleotide-free state is vital for elucidating its mechanical cycle.
• Switch I and switch II are key elements involved in nucleotide binding and release.

Purpose of the Study:

• To determine the structure of kinesin's switch I and switch II helix in the nucleotide-free state.
• To investigate the conformational changes of these elements upon microtubule binding.
• To elucidate the mechanism of microtubule activation and nucleotide ejection.

Main Methods:

• Cryo-electron microscopy (cryo-EM) of kinesin-decorated microtubules.
• Adaptation of single-particle reconstruction for analyzing decorated microtubules.
• High-resolution structural analysis of protein-microtubule interactions.

Main Results:

• Resolved the structure of switch I and switch II in the nucleotide-free state.
• Observed conformational changes in switch I and switch II relative to the microtubule-free state.
• Identified a potential role for switch I in adenosine diphosphate ejection.
• Revealed a microtubule-stabilized N-terminal extension of switch II, suggesting a signaling mechanism.

Conclusions:

• The conformational changes in switch I and switch II are critical for kinesin's motor function.
• Microtubule binding likely activates kinesin by stabilizing switch II, transmitting nucleotide state information.
• The adapted cryo-EM technique allows visualization of microtubule asymmetry and binding proteins at high resolution.