Reversible biological motion with unidirectional catalysis through inversion of ATPase orientation

Biological and engineered machines generally achieve reversibility through regulated switches in the directionality of fixed-orientation motors. Bacterial Tad pilus nanomachines extend and retract pili using a single motor with a unidirectional catalytic mechanism, a capability with no precedent. Using AlphaFold3 modeling, comparative structural analyses, and pilus activity assays, we find that the Tad motor ATPase CpaF achieves bidirectionality through physical inversion; alternating which face of the ATPase toroid engages the platform complex. The two orientations contact the pilus machinery in mutually exclusive extension- or retraction-specific configurations that drive reversible pilus dynamics using the same unidirectional catalytic cycle. Retraction requires conserved C-terminal residues in CpaF whose nucleotide-driven motions oppose those of the extension interface. Thus, nature has adopted a solution for reversible movement not yet conceived by human engineering.