Fig. 5: Tertiary contact formation monitored by FRET in space and time.

a Exon release is described by two FRET observables: dye distance and state dwell times. Intron–exon dissociation can occur from state 1 (Mg²⁺ free) or state 2 (Mg²⁺ bound) and is thus kinetically heterogeneous. b Occupation of Mg²⁺ binding sites as a function of the ion concentration using Mg²⁺ binding constants from NMR chemical shift perturbations (Eq. 5)¹⁵. Cartoons depict possible species with no (gray), one (dark green), or two Mg²⁺ bound (light green). The Mg²⁺ free species corresponds to the high FRET state 1 in a, while the other species together make up state 2. Diffuse Mg²⁺ ions are omitted for clarity. c Free energy diagram of tertiary contact formation featuring a reactant-like, unstructured transition state. Off-rates determine the stability of the interactions. Dotted black lines connect levels of the bound state at different metal ion concentrations. Source data are provided as a Source Data file.