FIGURE 1.

Single-molecule observation of DCCD inhibition of F₀F₁. A, experimental system for the observation of the F₀F₁ rotation. After the immobilization of purified F₀F₁ on a nickel-nitrilotriacetic acid (Ni-NTA)-coated glass, magnetic beads were attached by fusing the biotin-binding domain to the β subunit of F₀F₁. The rotation of the bead was observed using optical microscopy. The rotor (red) rotates counterclockwise against the stator (blue). In stalling experiments, the rotational angle of the bead was controlled by magnetic tweezers. B, time courses of the revolutions of rotating molecules before and after adding 1 mm DCCD. After 5 min of recording the rotation, a buffer containing 1 mm DCCD was infused into the chamber. When the buffer change was finished (∼1 min), the recording was restarted. Of the eight rotating molecules, six terminated their rotations, and the remaining two (red lines) kept rotating. Of the former six molecules, four had already terminated their rotation before restarting the recording, and the other two stopped after restarting the video recording.