Molecular Modeling Of Human Bitter Taste Receptors

Bitter taste perception is thought to evolve as a defense mechanism against ingesting poisonous substances because many toxins taste bitter. Although bitter taste receptors (T2Rs) have been definitely identified in humans and rodents since2000, little is currently known about the structure-function relationships of T2Rs due to the lack of3D structural information. Bitter taste receptors (T2Rs) belong to G-protein-coupled receptors (GPCRs). Due to inherent problems in the crystallization of GPCRs, structural determination of the members of this family has not been very successful. Therefore, computational protein modeling has been widely utilized to fill the gap. In this dissertation, computational methods were employed to elucidate the structure-function relationships of human bitter taste receptors (hTAS2Rs), especially the relationships of hTAS2R38and hTAS2R1.It has been suggested that sequence variants in hTAS2R38correlate with differences in bitterness recognition of phenylthiocarbamide (PTC). The three most common polymorphisms observed in TAS2R38occur at amino acid position49,262, and296, giving rise to two frequent haplotypes, PAV and AVI. hTAS2R38-PAV is sensitive to PTC while hTAS2R38-AVI has little or no sensitivity to PTC. Using the crystal structure of bovine rhodopsin as template, we generated the3D structure of hTAS2R38by comparative modeling. The PTC binding site in hTAS2R38was then predicted by molecular docking experiments. MD simulations of different hTAS2R38haplotypes in their free form and in complex with PTC were carried out in explicit lipid bilayer membranes. The predicted binding pocket is formed by Met100, Asn103and Gln104in TM Ⅲ, Phel97in TM Ⅴ, Phe264in TM Ⅵ, Met286TM Ⅶ and Met176in ECL2. P49V, A262V and V296I were found to locate in ICL1, TM Ⅵ and TM VII respectively. None of them has direct interactions with PTC. The MD simulation results showed that variants at position262and296might destroy the stability of TM VII due to the increase of hydrophobicity and steric hindrance.hTAS2Rl has been proved to be the receptor for bitter dipeptides. The structure of hTAS2R1was first modeled using many automated servers and meta-servers. The predictions from these servers were collected and consensus analysis was made to determine the right alignment between the target and the template. Then molecular docking experiments were carried out to identify the best model from the decoys with right sequence alignment and to generate the receptor-ligand complex models. Extensive all-atom molecular dynamics (MD) simulations of the selected best model in its free form and in complex with dipeptides in explicit lipid bilayer membranes were conducted to refine the models. Finally, the conformational changes induced by different ligands were also compared to gain insights into the activation mechanism of hTAS2Rl. The predicted binding pocket is formed by11residues from transmembrane helix Ⅲ (TM Ⅲ), TM Ⅴ, TM Ⅵ, TM Ⅶ and extracellular loop2(ECL2). Glu90in TM Ⅲ and Lys244in TM Ⅵ may be the most important two residues of the binding pocket. These two charged residues may be responsible for the dipeptide specificity of hTAS2Rl. Besides, the hydrophobic residues in the binding pocket may provide a hydrophobic surrounding for the side chains of dipeptides, which explains why there is an empirical correlation between the hydrophobicity of the peptides and their bitter taste. The intracellular loop II (ICL2) and TM III were found to play prominent roles in the process of activation. We proposed that a set of interactions between Phe115in ICL2and three residues (Tyr103, Lys106and Va1107) at the cytoplasmic end of TM III may serve as a conformational switch of hTAS2R1activation. These residues can hold the receptor in an inactive conformation through hydrophobic interactions, while the binding of bitter dipeptides can disrupt the hydrophobic interactions and thus open the conformational switch. The opening of the switch tends to cause ICL2and ICL3to move apart from each other, which may have direct implications in G-protein coupling. All of the residues involved in the switch are highly conserved among T2Rs. This indicates that the control switch we proposed may be universal in T2Rs.