Figure 3 | Scientific Reports

Figure 3

From: The Bitter Taste Receptor TAS2R16 Achieves High Specificity and Accommodates Diverse Glycoside Ligands by using a Two-faced Binding Pocket

Figure 3

Identification of ligand-specific structures. (a) Each of the four TAS2R16 ligands selected was tested against the entire TAS2R16 mutation library (573 individual mutations, n = 2–3) and values derived for each are compared. Concentrations chosen for mutation library screening were approximately 2 to 3 times higher than the EC50 value for each ligand to maximize the sensitivity of detecting decreases in signaling due to the mutation in each clone (3 mM salicin, 5 mM hexyl-β-glucoside, 1.6 mM 4-NP-β-mannoside, and 10 mM β-glucosaminide). Cutoffs of (100-3*SD), derived from wild-type positive control wells for each ligand, were used to identify clones that showed high Ca2+ flux for one ligand but low Ca2+ flux for the other (57% for salicin, 58% for hexyl-glucoside, 53% for 4-NP-β-mannoside, and 46% for β-glucosaminide). Of these critical residues, mutants that signaled 2.5-fold higher or lower were identified as the most significant (shown in red). Average values obtained from wild-type controls for each ligand are shown in blue. (b) Ca2+ flux activities are shown for clones expressing mutations that resulted in a ≥ 2.5-fold difference between Ca2+ flux activities for any two ligands.