Modulatory and catalytic modes of ATP binding by the calcium pump

We present crystal structures of the calcium-free E2 state of the sarcoplasmic reticulum Ca(2+)-ATPase, stabilized by the inhibitor thapsigargin and the ATP analog AMPPCP. The structures allow us to describe the ATP binding site in a modulatory mode uncoupled from the Asp351 phosphorylation site. The Glu439 side chain interacts with AMPPCP via an Mg(2+) ion in accordance with previous Fe(2+)-cleavage studies implicating this residue in the ATPase cycle and in magnesium binding. Functional data on Ca(2+) mediated activation indicate that the crystallized state represents an initial stage of ATP modulated deprotonation of E2, preceding the binding of Ca(2+) ions in the membrane from the cytoplasmic side. We propose a mechanism of Ca(2+) activation of phosphorylation leading directly from the compact E2-ATP form to the Ca(2)E1-ATP state. In addition, a role of Glu439 in ATP modulation of other steps of the functional cycle is suggested