Probing the dynamical interaction of the para-sulfonato-calix[4]arene with an antifungal protein

Calixarenes are hallmark molecules in supramolecular chemistry as guest cages for small ligands. They have also conversely proved their interest as auxiliary ligands toward assisted co-crystallization of proteins. These functionalized macromolecular cages target positively-charges residues, and notably surface-exposed lysines, with a site-selectivity finely characterized experimentally, but that remains to be assessed. Relying on a tailored molecular dynamics simulations protocol, we explore the association of para-sulfonato-calix[4]arenes with an antifungal protein, as a small yet most competitive system with 13 surface-exposed lysines. Our computational approach probe "de novo" the electrostatically-driven interaction, ruled out by a competition with salt bridges, not only corroborating the main binding site probed by X-ray, but also characterizing a second binding site that can act as a transient hub spot. The attach-pull-release (APR) method provides a very good assessment of the overall binding free energy measured experimentally (-6.42+-0.5 vs -5.45 kcal.mol-1 by isothermal titration calorimetry). This work also probes dynamical allosteric modifications upon ligand binding, and our computational protocol could be generalized to situate the supramolecular forces ruling out the calixarene-assisted co-crystallization of proteins.