Researchers at the University of North Carolina (UNC) at Chapel Hill School of Medicine have revealed the detailed structure of the bitter taste receptor, a protein called TAS2R14, and have shown where bitter-tasting substances bind to TAS2R14 and how they activate them, allowing us to taste bitter substances. This new, detailed information is important for discovering and designing drug candidates that can directly regulate taste receptors, with the potential to treat metabolic diseases such as obesity and diabetes, the team claimed.

“Scientists know very little about the structural make up of sweet, bitter, and umami taste receptors,” said Yoojoong Kim, PhD, a postdoctoral researcher in the lab of research lead Bryan Roth, MD, PhD, the Michael Hooker Distinguished Professor of Pharmacology. “Using a combination of biochemical and computational methods, we now know the structure of the bitter taste receptor TAS2R14 and the mechanisms that initializes the sensation of bitter taste in our tongues.”

Yoojoong Kim, PhD [UNC Department of Pharmacology]
Yoojoong Kim, PhD [UNC Department of Pharmacology]
The authors reported on their findings in Nature, in a paper titled, “Bitter taste receptor activation by cholesterol and an intracellular tastant.”

Humans can sense five different tastes: sour, sweet, umami, bitter, and salty, using specialized sensors on our tongues called taste receptors. Other than allowing us to enjoy delicious foods, the sensation of taste allows us to determine the chemical makeup of food and prevents us from consuming toxic substances. “As an initial defense system, bitter taste perception has a key role in sensing potentially toxic compounds via bitter taste receptors in the taste bud cells of the tongue,” the authors wrote.

For their newly reported work, the Roth lab researchers set out to address how exactly, we perceive bitter taste. TAS2R14s are members of the G protein-coupled receptor (GPCR) family of bitter taste receptors. TAS2R14 stands out from the other 26 members of the TAS2R family because it can identify dozens of substances known as bitter tastants. “Bitter taste sensing is mediated by type 2 taste receptors (TAS2Rs; also known as T2Rs), which represent a distinct class of G-protein-coupled receptors,” the authors further explained. “Among the 26 members of the TAS2Rs, TAS2R14 is highly expressed in extraoral tissues and mediates the responses to more than 100 structurally diverse tastants, although the molecular mechanisms for recognizing diverse chemicals and initiating cellular signaling are still poorly understood.”

The researchers aimed to elucidate the structural and functional mechanisms of the bitter taste receptor TAS2R14 both in relation to bitter taste sensing that occurs in native taste bud cells—this is the gustducin pathway—and to extraoral functions outside taste bud cells, for example, inhibitory G-protein pathways, they noted.

Through their studies, involving cryo-electron microscopy, biochemical, computational, and functional research, the team found that when bitter tastants come into contact with TAS2R14 receptors, the chemicals wedge themselves into to a specific spot on the receptor called an allosteric site, causing the protein to change its shape, and so activate the attached G protein.

Bryan Roth, MD, PhD [UNC Department of Pharmacology]
Bryan Roth, MD, PhD [UNC Department of Pharmacology]
This triggers a series of biochemical reactions within the taste receptor cell, leading to activation of the receptor, which can then send signals to tiny nerve fibers—through the cranial nerves in the face—to an area of the brain called the gustatory cortex. It is here where the brain processes and perceives the signals as bitterness. This complex signaling system occurs almost instantaneously.

While working to define its structure, researchers found another unique feature of TAS2R14—that cholesterol is giving it a helping hand in its activation. Prior research had made a connection between TAS2R14 activity and cholesterol, they noted. Bile acids, which are created in the liver, have similar chemical structures with cholesterol. Previous studies had in addition suggested that bile acids can bind and activate TAS2R14, but little is known about how and where they bind in the receptor.

“Recent evidence suggests that cholesterols and bile acids modulate several bitter taste receptors including TAS2R14,” the team noted. “However, the biological mechanisms by which TAS2R14 can be activated by cholesterols and bile acids remain unknown.” The new studies showed that cholesterol was residing in another binding site called the orthosteric pocket in TAS2R14, while the bitter tastant binds to the allosteric site, Kim said. “Through molecular dynamics simulations, we also found that the cholesterol puts the receptor in a semi-active state, so it can be easily activated by the bitter tastant.”

Representative cryo-EM maps (left) and models (right) for TAS2R14-Ggust-scFv16, colored according to each subunit, TAS2R14; green, Gαi1; purple, Gαgust; pink, Gβ1; sky blue, Gγ2;yellow, scFv16; gray.
Representative cryo-EM maps (left) and models (right) for TAS2R14-Ggust-scFv16, colored according to each subunit, TAS2R14; green, Gαi1; purple, Gαgust; pink, Gβ1; sky blue, Gγ2;yellow, scFv16; gray. [Kim et al (2024)]
Using their newfound structure data, the researchers found that bile acids might be binding to the same orthosteric pocket as cholesterol. While the exact role of bile acid or cholesterol in TAS2R14 remains unknown, it may play a role in the metabolism of these substances or in relation to metabolic disorders such as obesity or diabetes.

The authors summarized, “Our structures revealed two unexpected findings: (1) a cholesterol molecule in the presumed orthosteric binding site and (2) a novel intracellular positive allosteric modulator (PAM) site occupied by the bitter tastant compound 28.1 (cmpd28.1) … Our functional analysis identified cholesterol as an orthosteric agonist and the bitter tastant cmpd28.1 as a positive allosteric modulator with direct agonist activity at TAS2R14.” Cmpd28.1 is a recently developed potent agonist for TAS2R14 and a drug candidate that has been predicted to bind to a conventional class A orthostatic site, they wrote in their report. “Our findings provide insights into the bitter taste receptor TAS2R14 as a potential metabolite sensor that recognizes cholesterols and bile acids, with a novel intracellular PAM site for the recognition of bitter tastants.”

The discovery of this novel allosteric binding site for bitter-tasting substances is unique, the team claimed. The allosteric binding region is located between TAS2R14 and its coupled G protein is called G-protein alpha. This region is critical to form a signaling complex, which helps to transfer the signal from the taste receptor to the G-protein to the taste receptor cells.

“In the future, this structure will be key to discovering and designing drug candidates that can directly regulate G proteins through the allosteric sites,” noted Kim. “We also have the ability to affect specific G-protein subtypes, like G-protein alpha or G-protein beta, rather than other G-protein pathways that we don’t want to cause any other side effects.”

While running a genomics study, Kim and Roth found that the TAS2R14 protein complex is expressed outside the tongue, especially in the cerebellum in the brain, the thyroid, and the pancreas. Analysis of human tissues found TAS2R14 mRNA was highly expressed in different tissues. “We found higher expression of TAS2R14 mRNA in all the extraoral samples than in the tongue, with 100-fold higher expression in cerebellum, 50-fold higher in pancreas and small intestine, 26-fold higher in thymus, and 11-fold higher in adipose tissue compared with tongue.” An analysis of publicly available RNA-sequencing and single-cell RNA-seq datasets also confirmed TAS2R14 expression in multiple tissue types, indicating the highest expression levels in the cerebellum. “Collectively, these data suggest that TAS2R14 regulates cellular signaling, not in the tongue, but in various extraoral tissues, with particularly high expression in the cerebellum,” they wrote. Researchers are planning future studies to elucidate the function these proteins may have outside of the mouth.

Noting the need for further research, the authors concluded, “… our findings suggest that the bitter taste receptor TAS2R14, which is highly expressed in extraoral tissues, recognizes diverse compounds in terms of both number and structure, and may detect extracellular and intracellular metabolites as well as bitter tastants …”

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