The diffusive search mechanism of processive myosin class-V motor involves directional steps along actin subunits.

It is widely accepted that the vesicle-transporter myosin-V moves processively along F-actin with large steps of approximately 36 nm using a hand-over-hand mechanism. A key question is how does the rear head of two-headed myosin-V search for the forward actin target in the forward direction. Scanning probe nanometry was used to resolve this underlying search process, which was made possible by attaching the head to a relatively large probe. One-headed myosin-V undergoes directional diffusion with approximately 5.5 nm substeps to develop an average displacement of approximately 20 nm, which was independent of the neck length (2IQ and 6IQ motifs). Two-headed myosin-V showed several approximately 5.5 nm substeps within each processive approximately 36 nm step. These results suggest that the myosin-V head searches in the forward direction for the actin target using directional diffusion on the actin subunits according to a potential slope created along the actin helix.