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. 2013 May 31;8(5):e64675.
doi: 10.1371/journal.pone.0064675. Print 2013.

Coarse-grained/molecular mechanics of the TAS2R38 bitter taste receptor: experimentally-validated detailed structural prediction of agonist binding

Alessandro Marchiori  1 , Luciana Capece, Alejandro Giorgetti, Paolo Gasparini, Maik Behrens, Paolo Carloni, Wolfgang Meyerhof
Affiliations

Coarse-grained/molecular mechanics of the TAS2R38 bitter taste receptor: experimentally-validated detailed structural prediction of agonist binding

Alessandro Marchiori et al. PLoS One. .

Abstract

Bitter molecules in humans are detected by ∼25 G protein-coupled receptors (GPCRs). The lack of atomic resolution structure for any of them is complicating an in depth understanding of the molecular mechanisms underlying bitter taste perception. Here, we investigate the molecular determinants of the interaction of the TAS2R38 bitter taste receptor with its agonists phenylthiocarbamide (PTC) and propylthiouracil (PROP). We use the recently developed hybrid Molecular Mechanics/Coarse Grained (MM/CG) method tailored specifically for GPCRs. The method, through an extensive exploration of the conformational space in the binding pocket, allows the identification of several residues important for agonist binding that would have been very difficult to capture from the standard bioinformatics/docking approach. Our calculations suggest that both agonists bind to Asn103, Phe197, Phe264 and Trp201, whilst they do not interact with the so-called extra cellular loop 2, involved in cis-retinal binding in the GPCR rhodopsin. These predictions are consistent with data sets based on more than 20 site-directed mutagenesis and functional calcium imaging experiments of TAS2R38. The method could be readily used for other GPCRs for which experimental information is currently lacking.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Molecular Mechanics/Coarse-grained system set-up.
A) Schematic representation of the regions defined in the MM/CG model. The MM, I and CG regions are colored in green, orange, light blue, respectively. B) MM/CG representation of the hTAS2R38 receptor in complex with PTC. Water molecules and residues belonging to the MM and I regions are represented as lines. The agonist atoms are represented as orange spheres. The protein Cα atoms are represented in violet.
Figure 2
Figure 2. β2 AR in complex with the S-Car ligand.
A) S-Car binding site of the central structure of the only cluster of the MM/CG simulations. This cluster represent 100% of the conformations of the adduct. B) RMSD of the Cα atoms plotted as a function of time in the MM/CG simulations. C–D) Distribution of agonist-protein H-bonds in the MM/CG simulations of the β2 AR/S-Car complex based on the X-ray structure (black line) and on an homology model (violet line, this work).
Figure 3
Figure 3. Central structures of PTC/A - PROP/A and PTC/B - PROP/B main clusters emerging from our MM/CG simulations.
Residues forming mostly or exclusively hydrophobic contacts with the agonists (Met100, Phe264, Phe197, Trp99, Trp201, Tyr193) are colored in green, pink, orange, light violet, purple, yellow, respectively. The agonists are shown in ball-and-sticks representation and they are colored by atom type. The ECL2 loop interacts with the binding site only in the A complexes. It is shown in red cartoon. Selected distances for Asn179, Thr180, Arg181 and Asn183 residues in the ECL2 loop are shown for PTC/A and PROP/A.
Figure 4
Figure 4. Dose-response curves.
Dose-response curves of TAS2R38 wild type and mutants after stimulation with increasing PTC and PROP concentrations (0 to 1000 µM). Each point corresponds to the mean ± standard deviation. The mean is calculated from at least three independent experiments. A–C) PTC application, D–F) PROP application.
Figure 5
Figure 5. Dose-response curves.
Dose-response curves of TAS2R38 wild type and mutants after stimulation with increasing PTC and PROP concentrations (0 to 1000 µM). Each point corresponds to the mean ± standard deviation. The mean is calculated from at least three independent experiments. A–D) PTC application, E–H) PROP application.
Figure 6
Figure 6. Dose-response curves.
Dose-response curves of TAS2R38 wild type and mutants after stimulation with increasing PTC and PROP concentrations (0 to 1000 µM). Each point corresponds to the mean ± standard deviation. The mean is calculated from at least three independent experiments. A-E) PTC application, F-J) PROP application.
Figure 7
Figure 7. Agonists binding.
Binding of PTC and PROP to the TAS2R38 bitter receptor as emerging from MM/CG simulations and experiments. Residues forming hydrophobic interactions and H-bonds with the agonists are indicated in blue and red, respectively. Residues shaping the cavities are in black color. The ECL2 loop does not interact directly with the agonists.
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