Fig. 5. (a) Schematic representation of the competition experiment between GdC/AdU dinucleosides and aC/aU mononucleosides. Cyclic tetramers are in equilibrium with monomers (KT) and, under the conditions at which the competition experiments were carried out, such equilibrium is strongly (GdC) or moderately (AdU) shifted to the tetramer side. Upon addition of the complementary mononucleoside, two competing equilibria, both of them resulting in donor emission quenching due to energy transfer, would be established: (1) peripheral binding (Kp) to external sites of the cyclic tetramer (a hypothetical binding mode of aC to cGdC4 is shown) and (2) binding to the free dinucleoside monomer (Ka), which will shift the cyclotetramerization equilibrium toward the formation of GdC·aC/AdU·aU FRET pairs. (b and c) Normalized fluorescence emission changes (I/I0; λexc = 385 nm, T = 298 K, toluene) observed in the titration of (b) GdC with increasing amounts of aC ([GdC] = 1.0 × 10–4 M, [aC] = 2.0 × 10–4 M) and (c) AdU with increasing amounts of aU ([AdU] = 1.1 × 10–4 M, [aU] = 4.7 × 10–4 M). (d) Representation of the degree of cyclic tetramer association (χT) or emission changes (I/I0), as a function of the equivalents of complementary pyrimidine mononucleoside added for GdC + C (Fig. 4b; [GdC] = 1.0 × 10–4 M; red solid circles), AdU + U (Fig. 4c; [AdU] = 1.4 × 10–4 M; blue solid circles), GdC + aC (Fig. 5b; [GdC] = 1.0 × 10–4 M; red open circles), and AdU + aU (Fig. 5c; [AdU] = 1.1 × 10–4 M; blue open circles). (e) Normalized emission changes of GdC as a function of the equivalents of aC added at different GdC concentrations (see also Fig. S3C†).
