Figure 4.

Kinetic and affinity analysis of preQ₁ binding by the wildtype Tte riboswitch and position 14 variants. (A) Representative sensorgrams from surface plasmon resonance (SPR) showing preQ₁ association and dissociation to the wildtype riboswitch. Hydrogen-bonding diagrams for A14 are depicted for the effector-free state (binding pocket view) and preQ₁-bound state (L2 loop view) based on the crystal structures of this investigation (Figure 2A and B). PreQ₁ concentrations are shown in the key and colored lines represent background-subtracted data; black lines indicate the global fit to a 1:1 binding model. The chi² (RU²) for the fit was 1.8. The k_on and k_off rates and the apparent K_D values are in Table 2. (B) Representative sensorgrams for preQ₁ interaction with the A14(DAP) (2,6-diaminopurine) variant. The chi² (RU²) for the fit was 0.81. Putative hydrogen-bonding interactions for A14(DAP) are depicted for the effector-free state (binding pocket view) and preQ₁-bound state (L2 loop view) based on the wildtype crystal structures of this investigation. (C) Representative sensorgrams showing preQ₁ interactions with the A14(2AP) (2-aminopurine) variant. The chi² (RU²) for the fit was 1.5. Putative hydrogen-bonding interactions for A14(2AP) are depicted. (D) Representative sensorgrams showing preQ₁ interactions with the A14(Pur) (purine) variant. The chi² (RU²) for the fit was 0.85. Putative hydrogen-bonding interactions for A14(Pur) are depicted.