Supramolecular Host:Guest Arrays Site-Selectively Recognize Peptide Phosphorylation and Kinase Activity

A synergistic combination of cationic styrylpyridinium dyes and water-soluble deep cavitand hosts can recognize phosphorylated peptides with both site- and state-selectivity. Two mechanisms of interaction are dominant: either the cationic dye interacts with Trp residues in the peptide or the host:dye pair forms a heteroternary complex with the peptide, driven by both strong dye–peptide and cavitand–peptide binding (K d values up to 4 μM). The presence of multiple recognition mechanisms results in varying fluorescence responses dependent on the phosphorylation state and position, eliminating the need for covalent modification of the peptide target. Differential sensing aided by machine learning algorithms permits full discrimination between differently positioned serine phosphorylations with a minimal 3-component array. The array is fully functional in the presence of protein kinase A (PKA) and its required cofactors and capable of site-selective monitoring of serine phosphorylation at the privileged PKA motif, in the presence of serine residues that do not undergo reaction, illustrating the potential of the system in kinase-based drug screening.