Coupling of kinesin steps to ATP hydrolysis

Area of Science:

• Biochemistry
• Molecular Biology
• Cell Biology

Background:

• ATP-driven motor proteins like kinesin are crucial for intracellular transport.
• Understanding the precise mechanical steps and energy coupling of kinesin is essential for elucidating its function.
• Previous studies reported varied step sizes (5-8 nm) for kinesin, with analysis complicated by Brownian motion and rapid ATP turnover.

Purpose of the Study:

• To accurately quantify the step size of kinesin movement powered by a single ATP molecule.
• To resolve the relationship between ATP hydrolysis and kinesin's mechanical displacement.
• To determine if the observed coupling is consistent across different ATP concentrations.

Main Methods:

• Utilized advanced light microscopy with ultra-low positional drift (< 39 pms[-1]).
• Observed single kinesin molecules moving slowly at very low ATP concentrations (150nM) to separate temporal events.
• Applied a statistics-based analysis method accounting for unresolved movements to determine step size and ATP coupling.

Main Results:

• Kinesin exhibits a fundamental enzymatic cycle where a single ATP hydrolysis event is coupled to an 8.12 nm step distance, matching the microtubule protofilament lattice spacing.
• Movements of other step distances and complex ATP-to-step coupling ratios (e.g., 2 ATP per step) were excluded.
• The ratio of ATP consumption rate to stepping rate remained invariant across a broad range of ATP concentrations.

Conclusions:

• Kinesin's fundamental step size is 8 nm, directly coupled to the hydrolysis of one ATP molecule.
• This 1 ATP:8 nm coupling mechanism is a general feature of kinesin's enzymatic cycle, applicable from low to high ATP concentrations.
• The findings provide a clear quantitative understanding of kinesin's mechanical function at the molecular level.