Beschreibung:
<jats:p>The proton translocating ATP synthase is conceived as a rotatory molecular engine. ATP hydrolysis by its headpiece, CF<jats:sub>1</jats:sub>, drives the rotation of subunit γ relative to the hexagonally arranged large subunits, (αβ)<jats:sub>3</jats:sub>. We investigated transition states of the rotatory drive by polarized confocal fluorometry (POCOF) as applied to single molecules of engineered, immobilized and load‐free spinach‐CF<jats:sub>1</jats:sub>. We found that the hydrolysis of ATP caused the stepped and sequential progression of subunit γ through three discrete angular positions, with the transition states of γ being too shortlived for detection. We also observed the stepped motion of ϵ, whereas δ was immobile as (αβ)<jats:sub>3</jats:sub>.</jats:p>