Human annexin I is a member of the annexin family of calcium-dependent phospholipid binding proteins. The structure of an N-terminally truncated human annexin I (Δ-annexin I) and its interactions with Ca 2 + , Mg 2 + , and ATP were studied at the atomic level using nuclear magnetic resonance (NMR) spectroscopy. Since Δ-annexin I is a large protein, with a molecular weight of 35 kDa, a site-specific (carbonyl- 1 3 C, amide- 1 5 N) labeling technique was used to determine the interaction sites of Δ-annexin I with Ca 2 + , Mg 2 + , and ATP. The 1 3 C NMR study focused on the carbonyl carbon resonances of the histidine residues of Δ-annexin I. We found that ATP binds to Δ-annexin I, and that the ATP binding site is located in the 1-domain of annexin I. We also found that histidine-52 is involved in that site, and that the binding ratio of ATP to Δ-annexin I is 1:1.