The ATP-binding site of Ca2+-ATPase revealed by electron image analysis

The location of the ATP-binding site of a P-type ion pump, Ca2+-ATPase from rabbit sarcoplasmic reticulum, was examined by cryoelectron microscopy. A nonhydrolyzable analog of ATP, beta,gamma-bidentate chromium (III) complex of ATP (CrATP), was used to stabilize the enzyme in the Ca2+-occluded state. Tubular crystals were then induced by vanadate in the presence of EGTA, keeping CrATP bound to the enzyme. The three-dimensional structures of the crystals were determined at 14 Angstrom resolution by cryoelectron microscopy and helical image analysis. Statistical comparison of the structures with and without CrATP showed clear and significant differences at the groove proposed previously as the ATP-binding pocket.