Molecular Recognition and Fluorescence Sensing of Monophosphorylated Peptides in Aqueous Solution by Bis(zinc(II)−dipicolylamine)-Based Artificial Receptors

The phosphorylation of proteins represents a ubiquitous mechanism for the cellular signal control
of many different processes, and thus selective recognition and sensing of phosphorylated peptides and
proteins in aqueous solution should be regarded as important targets in the research field of molecular
recognition. We now describe the design of fluorescent chemosensors bearing two zinc ions coordinated
to distinct dipicolylamine (Dpa) sites. Fluorescence titration experiments show the selective and strong
binding toward phosphate derivatives in aqueous solution. On the basis of 1H NMR and 31P NMR studies,
and the single-crystal X-ray structural analysis, it is clear that two Zn(Dpa) units of the binuclear receptors
cooperatively act to bind a phosphate site of these derivatives. Good agreement of the binding affinity
estimated by isothermal titration calorimetry with fluorescence titration measurements revealed that these
two receptors can fluorometrically sense several phosphorylated peptides that have consensus sequences
modified with natural kinases. These chemosensors display the following significant features:  (i) clear
distinction between phosphorylated and nonphosphorylated peptides, (ii) sequence-dependent recognition,
and (iii) strong binding to a negatively charged phosphorylated peptide, all of which can be mainly ascribed
to coordination chemistry and electrostatic interactions between the receptors and the corresponding
peptides. Detailed titration experiments clarified that the phosphate anion-assisted coordination of the second
Zn(II) to the binuclear receptors is crucial for the fluorescence intensification upon binding to the
phosphorylated derivatives. In addition, it is demonstrated that the binuclear receptors can be useful for
the convenient fluorescent detection of a natural phosphatase (PTP1B) catalyzed dephosphorylation.