Fig 1.
Lactisole and 2,4-DP as sweet taste inhibitors.
(A) Structural formulas of (±)-lactisole and (±)-2,4-DP. (B) Dose-response curve showing the inhibition of the cellular responses of cells stably expressing the WT sweet taste receptor by (±)-lactisole and (±)-2,4-DP. Each inhibitor was added to 1 mM aspartame. The results for (±)-lactisole are shown in blue, and those for (±)-2,4-DP are shown in green. Data are shown as the mean ± SEM (n = 4). (C) Summary of mutational analysis for inhibitors (S.E.: standard error of each IC50). The results are summarized in S1 Table. Each ratio was calculated by dividing the value for the mutant by that for the WT. Mutants with IC50 ratios < 1/2 are colored in blue, ≥ 10 in orange, and ≥ 50 or N.D. in magenta, respectively. N.D., not determined since inhibitory activity was almost entirely eliminated.
Fig 2.
MD Simulations and their pre-simulations for lactisole and 2,4-DP.
(A) T1R3-TMD homology model created based on mGluR1 TMD (PDB ID: 4OR2). An enlarged view of the candidate ligand pocket is shown in the right window. Residues extracted by cellular analyses are indicated by colored stick illustrations. (B) Docking modes of (S)-lactisole (shown in blue) and (R)-lactisole (shown in pink) are shown in the left figure. (S)-2,4-DP (green) and (R)-2,4-DP (gray) are shown on the right. (C) MD minimizations (100 ns) were performed for each ligand from the docking models. The two left panels show the results for isomers of lactisole (upper panel: (S)-lactisole and lower panel: (R)-lactisole). The two right panels show the results for isomers of 2,4-DP (upper panel: (S)-2,4-DP and lower panel: (R)-2,4-DP). The estimated interactions between residues and the ligands are shown by dotted lines in the illustrations after MD: hydrogen bonds are shown in yellow, ionic bonds in magenta, and hydrophobic π-π interactions in cyan.
Fig 3.
Comparison of the results of MD minimizations for (S)-lactisole and (S)-2,4-DP.
(A) Results of the MD minimization of (S)-lactisole (shown in blue) and (S)-2,4-DP (shown in green). In the superimposed view after MD minimization, the (S)-lactisole model is shown by thin lines, and the (S)-2,4-DP model is shown by thick lines. (B) Van der Waals interaction areas between T1R3-TMD and (S)-lactisole or (S)-2,4-DP are shown.
Fig 4.
Sweet taste inhibitory activity of (S)-, (R)-lactisole and (S)-, (R)-2,4-DP.
(A) Flow chart of the synthesis of each isomer of lactisole and 2,4-DP. (B) Cellular response to 1 mM aspartame plus the indicated concentration of each compound against the cells expressing the WT sweet taste receptor. The left panel shows the results for the lactisole isomers: (S)-lactisole (blue) and (R)-lactisole (pink). The right panel shows the results for the 2,4-DP isomers: (S)-2,4-DP (green), and (R)-2,4-DP (gray). Data are shown as the mean ± SEM (n = 4).