Mitochondrial adenosine triphosphate (ATP) synthase comprises a membrane embedded F motor that rotates to drive ATP synthesis in the F subunit. We used single-particle cryo-electron microscopy (cryo-EM) to obtain structures of the full complex in a lipid bilayer in the absence or presence of the inhibitor oligomycin at 3.6- and 3.8-angstrom resolution， respectively. To limit conformational heterogeneity， we locked the rotor in a single conformation by fusing the F6 subunit of the stator with the δ subunit of the rotor. Assembly of the enzyme with the F6-δ fusion caused a twisting of the rotor and a 9° rotation of the F c-ring in the direction of ATP synthesis， relative to the structure of isolated F Our cryo-EM structures show how F and F are coupled， give insight into the proton translocation pathway， and show how oligomycin blocks ATP synthesis.