From Hydration Shell to Chelate Cavity: Funnel-Metadynamics Reveals the Sr2+ Chelation Pathway of Decorporol

The selective binding of Sr2+ over Ca2+ is very important for the development of chelating agents capable of selectively targeting radiostrontium in biological and environmental settings. Explicit-solvent molecular dynamics, Funnel-Metadynamics (Fun-MetaD), and density functional theory calculations were applied to resolve the molecular pathway of Ca2+ and Sr2+ binding by EDTA, Kryptofix 22, and Decorporol. EDTA was used as a benchmark system to validate the reparameterized force field protocol based on Minimal Basis Iterative Stockholder partial charges. The resulting relative free energy profiles reproduced the expected stronger stabilization of Ca2+ than Sr2+ by EDTA. Kryptofix 22 was included as an unfunctionalized diaza-18-crown-6 reference. Fun-MetaD profiles did not show a deeply stabilized contact-bound state. Higher concentration ITC measurements revealed weak, entropy-dominated Sr2+ association, while no reliable Ca2+ binding could be extracted under the same conditions. In contrast, simulations with Decorporol displayed a deeper contact-bound minimum for Sr2+, with minimum-to-bulk relative free energy differences of -113.2 ± 9.2 kJ/mol for Sr2+ compared with -98.2 ± 9.0 kJ/mol for Ca2+. Bound state integration and standard-state correction yielded binding free energies of -101.1 ± 2.1 kJ/mol for Sr2+ and -87.6 ± 6.4 kJ/mol for Ca2+, retaining the Sr2+-favoring trend. The simulations reveal that Decorporol binding proceeds through a two-step mechanism involving a solvent-shared ion-pair (SIP) intermediate followed by the formation of a deeply bound contact ion-pair (CIP) complex. The solvent-shared intermediate is not Sr2+-selective as both the Fun-MetaD profiles and density functional theory (DFT) calculations indicate similar Ca2+ and Sr2+ stability in this state. Selectivity emerges during the transition to the CIP complex, where the ion must undergo total dehydration to enter the Decorporol cavity. As Sr2+ has a more labile hydration shell, it can form the dehydrated contact-bound state more readily than Ca2+. Thus, Decorporol selectivity arises mainly from dehydration-gated stabilization of the CIP complex, rather than from initial ion recognition in the SIP state.