Determining the atomic charge of calcium ion requires the information of its coordination geometry in an EF-hand motif.

It is challenging to parameterize the force field for calcium ions (Ca ²⁺ ) in calcium-binding proteins because of their unique coordination chemistry that involves the surrounding atoms required for stability. In this work, we observed a wide variation in Ca ²⁺ binding loop conformations of the Ca ²⁺ -binding protein calmodulin, which adopts the most populated ternary structures determined from the molecular dynamics simulations, followed by ab initio quantum mechanical (QM) calculations on all 12 amino acids in the loop that coordinate Ca ²⁺ in aqueous solution. Ca ²⁺ charges were derived by fitting to the electrostatic potential in the context of a classical or polarizable force field (PFF). We discovered that the atomic radius of Ca ²⁺ in conventional force fields is too large for the QM calculation to capture the variation in the coordination geometry of Ca ²⁺ in its ionic form, leading to unphysical charges. Specifically, we found that the fitted atomic charges of Ca ²⁺ in the context of PFF depend on the coordinating geometry of electronegative atoms from the amino acids in the loop. Although nearby water molecules do not influence the atomic charge of Ca ²⁺ , they are crucial for compensating for the coordination of Ca ²⁺ due to the conformational flexibility in the EF-hand loop. Our method advances the development of force fields for metal ions and protein binding sites in dynamic environments.