pH-mediated fluorescence and G-quadruplex binding of amido phthalocyanines

A new family of G-quadruplex ligands termed "amido phthalocyanines" (APcs) was synthesized by reacting variable amino acids with tetraamino zinc phthalocyanine. Variation in the number of methylene units separating the APc scaffold from terminal ammonium groups systematically modulated ammonium pK(a) values that, in turn, mediated APc aggregation and DNA binding. Certain APcs exhibited nearly 1000-fold enhancements in fluorescence quantum yield upon binding G-quadruplex DNA under physiological conditions of pH and ionic strength. G-quadruplexes derived from the c-myc and c-kit promoters and the human telomeric repeat were evaluated for APc affinity and specificity using two complementary and direct fluorescence binding assays that revealed apparent dissociation constants ranging from 20 to 200 nM. Approximately 500-fold lower affinities for duplex and single-stranded DNAs were observed. Interestingly, APc-quadruplex binding was relatively insensitive to ionic strength (0.03-1 M KCl) but highly dependent on the pH of the solution. Our results provide a mechanism for the "turn-on" fluorescence properties exhibited by these compounds that will assist in future rational design of new G-quadruplex-specific fluorescent probes and drug candidates.