Toward the Rational Design of Tailored Ligands for Intrinsically Disordered Proteins through NMR

Abstract Understanding the driving forces governing the interactions between intrinsically disordered proteins and their potential ligands represents a key step toward the rational design of drugs targeting this type of proteins. Nuclear magnetic resonance spectroscopy provides a unique opportunity to achieve detailed atom-resolved information toward this ambitious goal. We present here a set of differently functionalized calix[4]arene-based multivalent ligands designed to target different regions of the well-characterized intrinsically disordered protein α-synuclein. Dedicated NMR experiments extended the analysis from backbone to side-chain nuclear spins, thus enabling the atom-resolved identification of short-range contacts between the calixarene ligands and methyl or aromatic groups of the protein surface, and revealing how these interactions refine the electrostatically driven association within different lysine-rich motifs. This approach provides a comprehensive description of how the macrocycles recognize and bind the target protein, offering a basis for the rational design of ligands capable of modulating protein function.