Biology subjects

Niv, M. Y.

Publications and source records attributed to Niv, M. Y..

13 recordsLinked to original sources

bioRxiv2026

Where do the ligands bind? Co-folding bitter taste GPCRs with the BitterDB chemical space

Bitterness is a key taste modality mediated in vertebrates by TAS2R G-protein-coupled receptors, which also function in diverse extraoral tissues. Recent cryo-EM structures have revealed a non-classical intracellular pocket in TAS2R14, raising the question of whether ligand pocket choice can be predicted computationally and what sequence features control it. Here we evaluate the Boltz-2 co-folding framework on all currently available agonist-TAS2R cryo-EM complexes and show that it correctly identifies the experimentally observed binding pocket for 12 of 15 pairs, including intracellular binding that docking into predicted receptor models fails to reproduce. Focusing on aristolochic acid, which binds intracellularly to TAS2R14 and extracellularly to TAS2R43, we use a series of in silico morphing experiments to pinpoint transmembrane helices 3 and 7, and specific residues within them, as key determinants of pocket preference. Extending the analysis to [~]1,500 agonist-receptor associations from BitterDB, we find that while most receptors are predicted to bind agonists predominantly in the extracellular pocket, several TAS2Rs may have both extracellularly and intracellularly binding ligands. Finally, by fine-tuning the Boltz-2 affinity module on [~]7,000 positive and negative experimental data points, we obtain a TAS2R-specific classifier that improves AUROC from 0.54 to 0.82 and average precision from 0.24 to 0.58 on a validation set.

biochemistry↗

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↗

bioRxiv2023

T1R3 subunit of the sweet taste receptor is activated by D2O in transmembrane domain-dependent manner

Deuterium oxide (D2O) is a water molecule in which both hydrogens are replaced by the heavier and rare isotope deuterium. We have previously shown that D2O has distinct sweet taste, which is mediated by the T1R2/T1R3 sweet taste receptor. Here we explore the effect of heavy water on T1R2 and T1R3 subunits. We show that D2O activates T1R3 transfected HEK293T cells similarly to T1R2/T1R3 transfected cells. The response to glucose dissolved in D2O is higher than to glucose dissolved in water. Mutations of Phenylalanine at position 7305.40 in the transmembrane domain of T1R3 to Alanine, Leucine or Tyrosine, impair or diminish activation by D2O, suggesting a critical role for T1R3 TMD domain in relaying the heavy water signal.

molecular biology↗

bioRxiv2022

Sensitivity of human sweet taste receptor subunits T1R2 and T1R3 to activation by glucose enantiomers

We have previously shown that L-glucose, the non-caloric enantiomer of D-glucose, activates the human sweet taste receptor T1R2/T1R3 transiently expressed in HEK293T cells. Here we show that D- and L-glucose can also activate T1R2 and T1R3 expressed without the counterpart monomer. Serine mutation to alanine in residue 147 in the binding site of T1R3 VFT domain, completely abolishes T1R3S147A activation by either L or D-glucose, while T1R2/T1R3S147A responds in the same way as T1R2 expressed without its counterpart. We further show that the original T1R2 reference sequence (NM_152232.1) is less sensitive by almost an order of magnitude than reference sequence at the time this study was performed (NM_152232.4). We find that out of the four differing positions, it is the R317G in the VFT domain of T1R2, that is responsible for this effect in-vitro. It is important both for practical assay sensitivity, and because glycine is found in this position in [~]20% of the world population. While the effects of the mutations and of the partial transfections were similar for D and L enantiomers, their dose response curves remained distinct, with L-glucose reaching an early plateau.

biochemistry↗

bioRxiv2022

Inhibiting a promiscuous GPCR: iterative discovery of bitter taste receptor ligands

The human GPCR family comprises circa 800 members, activated by hundreds of thousands of compounds. Bitter taste receptors, TAS2Rs, constitute a large and distinct subfamily, expressed orally and extra-orally and involved in physiological and pathological conditions. TAS2R14 is the most promiscuous member, with over 150 agonists and 3 antagonists known prior to this study. Due to the scarcity of inhibitors and to the importance of chemical probes for exploring TAS2R14 functions, we aimed to discover new ligands for this receptor, with emphasis on antagonists. To cope with the lack of experimental structure of the receptor, we used a mixed experimental/computational methodology which iteratively improved the performance of the predicted structure. The increasing number of active compounds, obtained here through experimental screening of FDA-approved drug library, and of chemically synthesized flufenamic acid derivatives, enabled the refinement of the binding pocket, which in turn improved the structure-based virtual screening reliability. This mixed approach led to the identification of 10 new antagonists and 200 new agonists of TAS2R14, illustrating the untapped potential of rigorous medicinal chemistry for TAS2Rs. The iterative framework suggested residues involved in the activation process, is suitable for expanding bitter and bitter-masking chemical space, and is applicable to other promiscuous GPCRs lacking experimental structures. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=172 SRC="FIGDIR/small/517821v4_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@30bd42org.highwire.dtl.DTLVardef@1d94fbforg.highwire.dtl.DTLVardef@2858borg.highwire.dtl.DTLVardef@18d9af2_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2022

Subjective assessment and taste strips testing of gustatory function, at home, and in the lab

Gustatory ability is an important marker of health status, including COVID-19 disease. We compare self-reporting with home and lab psychophysical "taste strips" tests in healthy subjects. The taste test consisted of paper strips impregnated with sweet, bitter, salty, or sour tastants, and with the trigeminal stimulus capsaicin, each in high and in low concentration. The test was carried out either in a controlled lab environment (74 participants, 47 women) with the strips being administered by the experimenter or self-administered by the participants at home (77 participants, 59 women). After self-reporting their subjective assessment of chemosensory ability, the participant identified the taste of each strip and rated intensity and pleasantness. Identification score, intensity, and pleasantness averaged over the 8 taste strips were similar between the lab and the home-administered tests. Self-rated taste ability did not correlate with any of these scores, but strongly correlated with self-rated smell ability in the lab group (r=0.73), and moderately correlated in the home group (r=0.51). Taste identification correlated with intensity ratings (r=0.63 lab, r=0.36 home) but not with the pleasantness ratings (r=-0.14 lab, r=0.1 home). The results of the taste strips test were similar in the lab and at home for healthy young participants and provide a baseline against which taste tests can be compared in future applications.

physiology↗

bioRxiv2022

BitterMatch: Recommendation systems for matching molecules with bitter taste receptors

Bitterness is an aversive cue elicited by thousands of chemically diverse compounds. Bitter taste may prevent consumption of foods and jeopardize drug compliance. The G protein-coupled receptors for bitter taste, TAS2Rs, have species-dependent number of subtypes and varying expression levels in extraoral tissues. Molecular recognition by TAS2R subtypes is physiologically important, and presents a challenging case study for ligand-receptor matchmaking. Inspired by hybrid recommendation systems, we developed a new set of similarity features, and created the BitterMatch algorithm that predicts associations of ligands to receptors with ~80% precision at ~50% recall. Associations for several compounds were tested in-vitro, resulting in 80% precision and 42% recall. The encouraging performance was achieved by including receptor properties and integrating experimentally determined ligand-receptor associations with chemical ligand-to-ligand similarities. BitterMatch can predict off-targets for bitter drugs, identify novel ligands and guide flavor design. Inclusion of neighbor-informed similarities improves as experimental data mounts, and provides a generalizable framework for molecule-biotarget matching.

biochemistry↗

bioRxiv2020

Sweet chirality: the taste of L and D glucose stereoisomers

Chirality plays a key role in biomolecular recognition. Typically, a change in chirality will dramatically affect ligand-receptor binding. However, both O_SCPLOWDC_SCPLOW-glucose and its enantiomer O_SCPLOWLC_SCPLOW-glucose elicit sweet taste in humans. We show that O_SCPLOWLC_SCPLOW- and O_SCPLOWDC_SCPLOW-glucose are perceived as similarly sweet by humans, and that in cell-based functional assays, both enantiomers activate the human sweet taste receptor TAS1R2/TAS1R3. We hypothesize that both O_SCPLOWLC_SCPLOW- and O_SCPLOWDC_SCPLOW-glucose occupy the orthosteric binding site in the VFT domain of TAS1R2. Using induced-fit docking to a homology model of this domain, we identify two subpockets in this binding site. The model suggests that glucose molecules can bind in either of these subpockets, which overlap with the predicted positions of monosaccharide units of sucrose. One subpocket is close to the hinge between the two lobes, and overlaps with aspartame and neotame site, the second subpocket overlaps with that described for sweetness enhancers. These findings suggest a framework for rational design of sweeteners combinations.

biochemistry↗

bioRxiv2020

Intense bitterness of molecules: machine learning for expediting drug discovery

Drug development is a long, expensive and multistage process geared to achieving safe drugs with high efficacy. A crucial prerequisite for completing the medication regimen for oral drugs, particularly for pediatric and geriatric populations, is achieving taste that does not hinder compliance. Currently, the aversive taste of drugs is tested in late stages of clinical trials. This can result in the need to reformulate, potentially resulting in the use of more animals for additional toxicity trials, increased financial costs and a delay in release to the market. Here we present BitterIntense, a machine learning tool that classifies molecules into "very bitter" or "not very bitter", based on their chemical structure. The model, trained on chemically diverse compounds, has above 80% accuracy on several test sets. BitterIntense suggests that intense bitterness does not correlate with toxicity and hepatotoxicity of drugs and that the prevalence of very bitter compounds among drugs is lower than among microbial compounds. BitterIntense allows quick and easy prediction of strong bitterness of compounds of interest for food and pharma industries. We estimate that implementation of BitterIntense or similar tools early in drug discovery and development process may lead to reduction in delays, in animal use and in overall financial burden. Significance StatementDrug development integrates increasingly sophisticated technologies, but extreme bitterness of drugs remains a poorly addressed cause of medicine regimen incompletion. Reformulating the drug can result in delays in the development of a potential medicine, increasing the lead time to the patients. It might also require the use of extra animals in toxicity trials and lead to increased costs for pharma companies. We have developed a computational predictor for intense bitterness, that has above 80% accuracy. Applying the classifier to annotated datasets suggests that intense bitterness does not correlate with toxicity and hepatotoxicity of drugs. BitterIntense can be used in the early stages of drug development to identify drug candidates that require bitterness masking, and thus reduce animal use, time and monetary loss.

pharmacology and toxicology↗

bioRxiv2020

Sweet taste of heavy water

Hydrogen to deuterium isotopic substitution has only a minor effect on physical and chemical properties of water and, as such, is not supposed to influence its neutral taste. Here we conclusively demonstrate that humans are, nevertheless, able to distinguish D2O from H2O by taste. Indeed, highly purified heavy water has a distinctly sweeter taste than same-purity normal water and adds to perceived sweetness of sweeteners. In contrast, mice do not prefer D2O over H2O, indicating that they are not likely to perceive heavy water as sweet. HEK 293T cells transfected with the TAS1R2/TAS1R3 heterodimer and chimeric G-proteins are activated by D2O but not by H2O. Lactisole, which is a known sweetness inhibitor acting via the TAS1R3 monomer of the TAS1R2/TAS1R3, suppresses the sweetness of D2O in human sensory tests, as well as the calcium release elicited by D2O in sweet taste receptor-expressing cells. The present multifaceted experimental study, complemented by homology modelling and molecular dynamics simulations, resolves a long-standing controversy about the taste of heavy water, shows that its sweet taste is mediated by the human TAS1R2/TAS1R3 taste receptor, and opens way to future studies of the detailed mechanism of action. One sentence summaryHeavy water elicits sweet taste for humans via the TAS1R2/TAS1R3 taste receptor.

biochemistry↗