A chimeric kinesin-1 head/kinesin-5 tail motor switches between diffusive and processive motility

Area of Science:

• Cell Biology
• Molecular Motors
• Biophysics

Background:

• Kinesin-5 motors are homotetrameric proteins essential for mitotic spindle assembly and chromosome segregation.
• These motors slide antiparallel microtubules (MTs) apart, a process requiring precise regulation during mitosis.
• The mechanism of allosteric communication between kinesin-5 motor domains, potentially involving tail domains, is not fully understood.

Purpose of the Study:

• To investigate the regulatory mechanisms of kinesin-5 motors using a novel chimera.
• To elucidate the distinct motile states and transitions involved in kinesin-5 function.
• To explore the role of tail domains in kinesin-5 motor regulation.

Main Methods:

• Single-molecule fluorescence microscopy was employed to study a kinesin-1 head/kinesin-5 tail chimera (DK4mer).
• Motility assays were performed on single and antiparallel microtubules under varying buffer ionic strengths.
• Transition rates between distinct motile states and unbinding rates were measured.

Main Results:

• DK4mer exhibited processive motility on single MTs, interspersed with pauses, and diffused with ADP.
• DK4mer actively slid antiparallel MTs apart in the presence of ATP, similar to native Eg5.
• Diffusive and processive states were clearly distinguishable, allowing quantification of transition and unbinding rates, revealing two MT interaction modes separated by an energy barrier.

Conclusions:

• A two-state model for kinesin-5 motor interaction with microtubules is proposed, involving switching between bound states.
• This switching mechanism is likely allosterically controlled by the opposing tetramer end, providing a regulatory scheme.
• The findings are relevant for understanding the regulation of native kinesin-5 motors in mitosis.