Fig. 2

Trp43 conformational exchange does not follow Arrhenius behavior in DANCERs. a ¹H-¹⁵N HSQC ZZ-exchange crosspeak intensity curves for the W43ε minor to major state transition show that this transition slows in DANCER-3 as temperature increases. This unusual behavior is not observed with T17, demonstrating a different mode of exchange for W43ε than monomer-dimer exchange. b Arrhenius plots demonstrate Arrhenius behavior for both W43ε and T17 exchange profiles in WT-A34F, as demonstrated by similar slopes (i.e., activation energies) for all four transitions. DANCER-3 however exhibits non-Arrhenius behavior, as shown by a W43ε minor to major state transition with a non-physical negative activation energy, which indicates that this residue does not follow a two-state Arrhenius model. In contrast, a normal Arrhenius behavior is observed for DANCER-3 T17. c In DANCERs, non-Arrhenius behavior for W43ε transitions is observed due to an altered monomer-dimer equilibrium. As dimerization of DANCERs is favored at high temperature and W43 exchange is inhibited in the dimer, observed rates (k_obs) of W43 exchange are reduced when temperature increases. In contrast, k_obs for exchange of the T17 residue found at the dimer interface increase with temperature, indicating that dimerization kinetics follow Arrhenius behavior. Non-Arrhenius behavior for W43ε transitions was not detected in Gβ1 variants that do not undergo W43 conformational exchange, suggesting that all kinetics measured for these variants report only on dimerization.