Structure and Stabilization Mechanism of ATP^4–·Mg^2+2–: a Joint Negative Ion Photoelectron Spectroscopic and Computational Study^†

MgATP is a stable complex formed by the chelation of Mg²⁺ with deprotonated adenosine-5'-triphosphate (ATP). In the cellular environment, MgATP plays a critical role in ATP hydrolysis, releasing substantial energy to support essential biological functions. To understand the structure and stabilization mechanism of MgATP, we conducted a joint negative ion photoelectron spectroscopic and computational study of the ATP^4–·Mg^2+2– complex dianion, using ATP^4–·2H^+2– as a reference. The experimentally determined adiabatic and vertical detachment energies (ADE and VDE) of ATP^4–·Mg^2+2– at 20 K are 3.51 ± 0.05 eV and 3.82 ± 0.05 eV, respectively. The major spectral features of ATP^4–·Mg^2+2– are attributed to two theoretically identified isomers with unfolded geometries, which are stabilized primarily by electrostatic interactions between Mg²⁺ and the triphosphate and ribose groups, with four deprotonated oxygens forming a pseudo-tetrahedral coordination. In contrast, ATP^4–·2H^+2– exhibits a fundamentally different stabilization mechanism. Although most of the fifteen identified ATP^4–·2H^+2– isomers also adopt unfolded geometries, they are primarily stabilized by intramolecular hydrogen bonds within the triphosphate group and between triphosphate and ribose groups. The interaction between ATP^4– and two protons is found to be much weaker than that with Mg²⁺, and ATP^4–·2H^+2– exhibits substantial structural flexibility compared to ATP^4–·Mg^2+2– due to the conformational constraint of the triphosphate chain by Mg²⁺. Thirteen ATP^4–·2H^+2– isomers with unfolded geometries likely account for the major high-EBE (electron-binding-energy) spectral features. Notably, for the first time, a low EBE and temperature-dependent spectral feature is observed and attributed to two folded isomers of ATP^4–·2H^+2–, which exist at 20 K but disappear at room temperature. This study provides valuable molecular-level insights into cellular MgATP that resides within the hydrophobic pockets of proteins.