F(1)-ATPase is the smallest mechanical motor known. Tentoxin, a cyclic peptide produced by phytopathogenic fungi, inactivates the F(1) motor in sensitive plants at nanomolar to micromolar concentrations, whereas higher concentrations surpass the natural activity of the enzyme. Single molecule studies now have clarified the molecular steps involved in both processes. Inactivation delays the dwell time of a single step in the complete 360 turn and results in an asymmetric rotation of the central rotor subunit. In contrast, rotation in the stimulated F(1) particle is smooth and accompanied by strongly reduced ADP inhibition. Our study provides for the first time the direct observation of a noncompetitively inhibited state of the enzyme and directly visualizes the regulation of the molecular motor by an external natural compound. In addition, the ADP release step during catalysis was revealed by analysis of the single molecule rotation behavior. Hence, tentoxin is a sophisticated molecular tool to mark and control certain catalytic steps within the reaction pathway of the molecular F(1) motor.