All-or-None Selectivity in Probing Polarity-Determined Trinucleotide Repeat Foldings with a Parity Resolution by a Beyond-Size-Matching Ligand

Abnormal amplification of trinucleotide repeats (TNRs) is associated with neurodegenerative diseases by forming a particular hairpin bulge. It is well known that the polarity and parity of TNRs can regulate the formed hairpin structures. Therefore, there is a great challenge to efficiently discriminate the hairpin structures of TNRs with substantial selectivity. Herein, we developed a fluorescent ligand of pseudohypericin (Pse) with a beyond-size-matching (BSM) geometry to selectively sense hairpin structures of GTC and CTG TNRs. The GTC hairpin structures can bind with Pse dominantly at extreme T–T mismatches by the virtue of their most extrahelical conformations, while there is no binding event to occur with the polarity-inverted counterpart CTG hairpin structures because of the limited space provided by their intrahelical T–T mismatches. In addition, this all-or-none response with the polarity-dependent folding (PoDF) is independent of the length of these TNRs. Interestingly, the parity-dependent folding (PaDF) of GTC hairpin structures can also be resolved. Besides pure TNRs, the competency of this BSM ligand to sense the PoDF and PaDF effects was also generalized to DNAs with TNRs occurring at loop and stem end regions. To our knowledge, this is the first experimental observation with the state-of-the-art performance over the fluorescence measurement of PoDF and PaDF in TNRs. Our work provides an expedient way to elucidate the TNR folding by designing ligands having BSM features.