To study domain organization and movements in the reaction cycle of heavy metal ion pumps, CopA, a bacterial Cu+-ATPase from Thermotoga maritima was cloned, overexpressed, and purified, and then subjected to limited proteolysis using papain. Stable analogs of intermediate states were generated using AMPPCP as a nonhydrolyzable ATP analog and AlFx as a phosphate analog, following conditions established for Ca2+-ATPase (SERCA1). Characteristic digestion patterns obtained for different analog intermediates show that CopA undergoes domain rearrangements very similar to those of SERCA1. Digestion sites were identified on the loops connecting the A-domain and the transmembrane helices M2 and M3 as well as on that connecting the N-terminal metal binding domain (NMBD) and the first transmembrane helix, Ma. These digestion sites were protected in the E1P.ADP and E2P analogs, whereas the M2-A-domain loop was cleaved specifically in the absence of ions to be transported, just as in SERCA1. ATPase activity was lost when the link between the NMBD and the transmembrane domain was cleaved, indicating that the NMBD plays a critical role in ATP hydrolysis in T. maritima CopA. The change in susceptibility of the loop between the NMBD and Ma helix provides evidence that the NMBD is associated to the A-domain and recruited into domain rearrangements and that the Ma helix is the counterpart of the M1 helix in SERCA1 and Mb and Mc are uniquely inserted before M2.