Motor domain-based motility system and motile properties of alpha heavy chain in Tetrahymena outer arm dynein

• A motor domain-based motility system was established in Tetrahymena outer arm dynein.
• The functional motor domain of DYH3 was purified using an in situ digestion method.
• The DYH3 motor domain exhibited ATP concentration-dependent inhibition.
• The DYH3 tail was not essential for ATP concentration-dependent inhibition.

Axonemal dynein plays an essential role in ciliary motility, and impaired ciliary motility causes human diseases such as primary ciliary dyskinesia (PCD). The motor domain of axonemal dynein powers ciliary motility and its function is regulated by several accessary proteins bound to the tail region. Therefore, to understand the essential properties of dynein motility, examining the motile properties of the motor domain without the tail is necessary. In this study, the functional motor domain of the alpha heavy chain in Tetrahymena outer arm dynein was purified, and its motile properties were examined using an in vitro motility system. The purified protein caused microtubules to glide at a velocity of 5.0 μm/s with their minus-end trailing, and motility was inhibited in an ATP concentration-dependent manner, which is in contrast with kinesin1. This method could be applicable to other axonemal dyneins and will enable further molecular studies on diverse axonemal dyneins and ciliary motility.