Biology subjects

Rainish, A.

Publications and source records attributed to Rainish, A..

4 recordsLinked to original sources

bioRxiv2026

Pharmaceutical TAS2R14 Agonists Display Diverse Potency, Efficacy, and Binding-Site Sensitivity

Bitter taste receptors (TAS2Rs) are G-protein coupled receptors that detect chemically diverse compounds, including many clinically used drugs. TAS2R14 is expressed in many extraoral tissues and is activated by hundreds of ligands, including pharmaceutical drugs. Recent cryo-EM structures revealed a previously unrecognized intracellular binding pocket in TAS2R14, raising new questions regarding ligand binding modes. Here, we investigated the activation of TAS2R14 by Tamoxifen, Carbimazole, and Lidocaine using cell-based assays measuring proximal G-protein recruitment (BRET2) and downstream signaling (IP-One). Tamoxifen and Carbimazole activated TAS2R14 with EC50 values in the low micromolar range, whereas Lidocaine required substantially higher concentrations. Targeted receptor mutations were used to evaluate the contribution of extracellular and intracellular binding regions to agonist activity. Carbimazole and Lidocaine showed greater dependence on the intracellular and extracellular positions, respectively, while Tamoxifen displayed assay-dependent, but overall modest sensitivity to the tested mutations. Thus, although existing drugs can activate TAS2R14 through distinct binding modes, TAS2R14-directed repurposing will depend on whether effective local receptor concentrations can be achieved through appropriate delivery strategies.

pharmacology and toxicology↗

bioRxiv2025

Intracellular and Dual-Site Inhibition of a bitter taste GPCR

Bitter taste receptors (TAS2Rs) are G protein-coupled receptors expressed in both gustatory and extraoral tissues and activated by a broad range of compounds. TAS2R14 is among the most promiscuous members of this family, responding to many structurally diverse ligands. Cryo-electron microscopy structures of TAS2R14 have revealed agonists binding in an intracellular pocket, raising the question of the main sites of interaction for known TAS2R14 antagonists. To address this, we examined the effects of mutations at residues located in the extracellular and intracellular regions on receptor inhibition by three antagonist compounds. Mutations in the extracellular region reduced the inhibitory effect of LF22, whereas all three compounds showed reduced inhibition in the intracellular mutants. Computational co-folding of these ligands with TAS2R14 supported these observations, indicating that LF22 interacts with both top and bottom binding sites, whereas LF1 and probenecid engage predominantly the intracellular site adjacent to the G protein interface. Interestingly, LF1 is much more potent for TAS2R16 than its known inhibitor probenecid. These findings reveal distinct inhibitory mechanisms among TAS2R antagonists and provide new insights towards designing inhibitors of bitter taste.

biochemistry↗

bioRxiv2025

Dynamics of Ligand Binding Sites and Chloride Penetration in a Bitter Taste GPCR

Taste perception strongly influences food choice, drug compliance, and dietary behavior. Bitter taste receptors belong to a subfamily of Family A GPCRs, while having unique features and deviations from conserved motifs. Recent CryoEM structures revealed that the human bitter taste receptor TAS2R14 contains not only the canonical extracellular binding site, but also a novel intracellular site, raising questions about their interplay. Using molecular dynamics simulations, we examined how binding of aristolochic acid in the intracellular site and cholesterol at the extracellular site, alone or together, affects receptor dynamics and pocket communication. We observed that cholesterol binding in the extracellular site expanded the intracellular pocket volume and induced conformational changes in TM6, whereas aristolochic acid binding in the intracellular site had no such effect on the extracellular site. Notably, in cholesterol-only simulations, a chloride ion entered from the intracellular side and interacted with Arginine 55 (BW position 2.50), revealing an inverted ion-binding pattern to Family A GPCRs, where aspartate 2.50 is known to bind a sodium ion to stabilize an inactive state. Our results suggest chloride-dependent and cholesterol-dependent modulation mechanisms in the dual-site dynamics of TAS2R14.

bioinformatics↗

bioRxiv2024

Intracellular binding pocket revealed in the human bitter taste receptor TAS2R14

Bitter taste receptors (TAS2Rs), a subfamily of G-protein coupled receptors (GPCRs) expressed orally and extraorally, elicit signaling in response to a large set of ligands. Among the 25 functional TAS2Rs encoded in the human genome, TAS2R14 is the most promiscuous, and responds to hundreds of chemically diverse agonists. Here, we present the cryo-electron microscopy (cryo-EM) structure of the human TAS2R14 (hTAS2R14) in complex with its cognate signaling partner gustducin, and bound to flufenamic acid (FFA), a clinically approved nonsteroidal anti-inflammatory drug. The structure reveals an unusual binding mode for FFA, where two copies are bound at distinct binding pockets: one at the canonical GPCR site within the trans-membrane bundle, and the other in the intracellular facet, bridging the receptor with gustducin. Combined with site-directed mutagenesis and the design of a fluorescent FFA derivative for pocket-specific ligand binding BRET assays, our studies support a dual binding mode for FFA in TAS2R14. These results fill a gap in the understanding of bitter taste signaling and provide tools for guided design of TAS2R-targeted compounds.

biochemistry↗