Figure 5: Structural dynamics of ECL3 in OR function.

a) Residue R262^6x59 in ECL3 makes a critical contact with propionate. Residue Q181^45x53 in ECL2 is highlighted. b) Molecular dynamics simulations of OR51E2 with propionate removed shows increased flexibility of R262^6x59. Representative snapshots are displayed from five independent simulation replicates after 1000 ns of simulation time. c) In simulations of wild-type (WT) OR51E2 bound to propionate, the minimum distance between R262^6x59 and G198^5x39 heavy atoms is stable and similar to the cryo-EM structure. Simulations of WT OR51E2 without propionate (no ligand) show increased minimum distance between R262^6x59 and G198^5x39. In simulations of Q181^45x53D mutant without propionate, the minimum distance between R262^6x59 and G198^5x39 is similar to WT OR51E2 bound to propionate. Minimum distance was measured between R262^6x59 sidechain atoms and G198^5x39 main chain atoms (excluding the hydrogens) over the course of 1000 ns MD simulation (see Extended Data Fig. 8 for simulation replicates). Thick traces represent smoothed values with an averaging window of 8 nanoseconds; thin traces represent unsmoothed values. d) Conservative mutagenesis of Q181^45x53 shows that the Q181^45x53D mutant is constitutively active, potentially because it substitutes a carboxylic acid in the OR51E2 binding pocket. e) Comparison of cryo-EM structure of OR51E2 with the AlphaFold2 predicted structure shows high similarity in the extracellular domain with the exception of the ECL3 region. The AlphaFold2 model shows an outward displacement of R262^6x59 and ECL3 similar to simulations of apo OR51E2. f) AlphaFold2 predictions for all human odorant receptors show low confidence in the ECL3 region and high confidence in other extracellular loops. g) A model for ECL3 as a key site for odorant receptor activation.