MX2007016411A - Identification of human t2r recepors that are activated by bitter molecules in coffee (chlorogenic lactones) and related assays for identifying human bitter taste modulators - Google Patents

Abstract

The present invention relates to the discovery that specific human taste receptors in the T2R taste receptor family respond to particular bitter compounds, i.e., chlorogenic lactone compounds that contribute at least partially to the bitter taste of many coffee beverages. The present invention further relates to the use of these receptors in assays for identifying ligands that modulate the activation of these taste receptors by chlorogenic lactones and related compounds and which may be used as additives and/or removed fromfoods, beverages and medicinals in order to modify (block) T2R-associated bitter taste. A preferred embodiment is the use of the identified compounds as additives in coffee and coffee-flavored foods, beverages and medicinals.

Description

IDENTIFICATION OF HUMAN T2R RECEIVERS THAT ARE ACTIVATED BY AMARGAS MOLECULES IN COFFEE (CHLOROGENIC LACTONS) AND RELATED TESTS TO IDENTIFY TASTE MODULATORS HUMAN AMARGO FIELD OF THE INVENTION The present invention relates to the elucidation of bitter compounds that activate a number of previously reported human G protein-coupled receptors (GPCRs) in the T2R family that are involved in the perception of bitter taste. Specifically, the invention involves the discovery that hT2R8, hT2R14 and hT2R54 respond specifically to chlorogenic lactones that are responsible at least in part for the bitter taste of coffee. Therefore, the subject T2Rs can be used to identify compounds that modulate, preferably block, the bitter taste, for example, of bitter compounds found in coffee and related bitter tastes. More specifically, the present findings indicate that subject human taste receptors, fragments, variants or chimeras thereof, including orthologs, splice variants, single nucleotide polymorphisms (SNPS), and genetically engineered mutants thereof, are useful in assays, preferably high-throughput cell-based assays, to identify compounds that modulate (preferably block) the bitter taste of the chlorogenic lactone molecules, as well as structurally related compounds and other compounds that activate these receptors. The compounds identified that use these assays can be used as additives in foods, beverages or medicinal products to improve their taste. Additionally, the invention relates to modified foods, beverages and medicinal substances that are treated and formulated in order to reduce or eliminate the bitter compounds that activate the subject T2Rs. DESCRIPTION OF THE RELATED TECHNIQUE One of the basic taste modalities that humans can recognize is the bitter taste. The physiology of bitter taste until very recently was poorly understood. Recent studies have begun to illuminate flavor biology (Lmdemann, Nature (2001)). It is now believed that many bitter compounds produce bitter taste by interacting with cell surface receptors. These receptors belong to the family of seven transmembrane receptors that interact with intracellular G proteins. A novel family of GPCRs, called T2Rs, has been identified in humans and rodents (Adler et al., Cell 100 (6): 693-702 (2000); Chandrashekar et al., Cell 100 (6): 703-711 (2000); Matsunami H, Montmayeur JP, Buck L B. Nature 404 (6778): 601-4 (2000)). Several lines of evidence suggest that T2Rs mediate responses to bitter compounds. First, the genes, T2R, are specifically expressed in a subset of flavor receptor cells of the tongue and palatal epithelia. Second, the gene for one of the human T2Rs (hT2Rl) is located at a chromosomal site that binds to the sensitivity of the bitter compound 6-n-propyl-2-thiouracil in humans (Adler et al., (Id.) (2000). )). Third, one of the mouse T2Rs (mT2R5) is located at the chromosomal site that binds to the bitter compound cycloheximide sensitivity in mice. It was also shown that mT2R5 can activate gustducin, G protein specifically expressed in flavor cells and linked to the transduction of bitter stimuli (ong et al., Nature 381: 796-800 (1996)). Activation of gustducin by mT2R5 occurs only in response to cycloheximide (Chandrashekar et al., (Id.) (2000). Thus, it has been proposed that the mT2R family mediates the bitter taste response in mice, whereas the hT2R family mediates the bitter taste response in humans Several T2Rs have also been identified as receptors by certain ligands (Chandrashekar et al., (Id.) 2000; Bufe et al., (Id.) 2002; Kim et al. Science 299, 2003; Pronin et al., Chemical Senses 29, 2004; Behrens et al., BBRC 319, 2004; Kuhn et al., J. Neuroscience 24, 2004; Bufe et al., Current Biology, 15, 2005). It has also been suggested that each hT2R is capable of binding multiple bitter ligands. This hypothesis is based on the fact that the hT2R family consists of only 24 identified members while humans can recognize hundreds of different compounds as bitter. Sequences of hT2Rs have been previously reported and disclosed in the PCT applications published by Zuker et al. (WO 01/18050 A2, (2001)) and Adler et al. (WO 01/77676 Al (2001)) both of which are incorporated by reference in its entirety herein. One of the difficulties in studying the function of T2R is that these receptors are not easily expressed in cell lines of cultured mammals. To improve the expression of T2R a well-expressed N-terminal GPCR sequence, rhodopsin, was bound to T2R sequences (Chandrashekar et al., (Id.) 2000). This N-terminal marking also allows the ease of monitoring the expression of protein due to a free antibody. While incorporation of the rhodopsin tag improves the expression of some T2Rs in mammalian cell lines, many of them are still not expressed well enough for functional studies. In a different procedure mT2R5 was successfully expressed in Sf9 insect cells and used for functional studies using the biochemical GTP binding assay (Chandrashekar et al., (Id.) 2000). In the previous patent application of the Requesters, the document of the U.S. Serial No. 10 / 191,058, incorporated by reference herein, applicants discovered ligands that specifically activate different human T2Rs. Additionally, the applicants presented the provisional document of U.S. Serial No. 60 / 650,555 on February 8, 2005, which additionally identifies bitter ligands that include acetaminophen, ranitidine, stpgnin and denatonium that specifically bind to seven specific human T2Rs, and related assays. However, although it has been reported and the understanding that T2R members regulate bitter taste, there is a need for the identification of specific ligands that activate specific T2R receptors. A greater understanding of the binding properties of the different T2Rs, particularly human T2Rs, would be highly beneficial since they greatly facilitate the use thereof in the selection of compounds that have desired flavor modulating properties, i.e., that block or inhibit the taste of the specific bitter compounds. Additionally, of particular relevance to the present invention, there is a particular need to identify compounds that can be used to inhibit the bitter taste associated with coffee. While coffee consumption in the United States and around the world has increased substantially through recent years, a prevalent complaint of many coffee drinkers is that many coffees induce a bitter after taste. Therefore, assays with the identification of compounds that cause coffee to exhibit a bitter after taste and / or which identify compounds that block the bitter after taste of coffee would be beneficial towards the production of coffees having improved acceptability. With respect thereto, it has been reported that roasting and processing coffee has an effect on the formation of lactones of chlorogenic acid in coffee (Furah et al., J. Agrie, Food Chem. 53 (5): 1505-13 ( 2005), Variyar et al, J. Agrie, Food Chem. 51 (27): 7495-50 (2003)). BRIEF DESCRIPTION OF THE INVENTION To that end, the present invention relates to the discovery that several taste receptors in the T2R family, particularly hT2R8, hT2R14 and hT2R54 respond specifically to the chlorogenic lactones found in coffee that are hypothesized to be responsible, at least in part, for the bitter taste of coffee beverages. These findings were made using cell-based assays that measured the activity of the T2Rs using cells expressing a particular T2R in the presence and absence of the specific bitter ligands. In particular, as described in more detail below, the HEK cell lines expressing the specific T2Rs identified above on their surface and which also express a chimeric G protein that functionally couples to the T2Rs were used in cell-based assays that detected changes in intracellular calcium concentrations, and were found to be specifically activated by specific bitter compounds (a number of chlorogenic lactone molecules) found in coffee and other foods and beverages while other hT2Rs were not activated under similar conditions. Therefore, the invention encompasses the use of these human taste receptors in assays, preferably in high throughput assays, to identify compounds that modulate, preferably block the activation of these receptors by chlorogenic lactones, derivatives thereof and other bitter compounds . Also, the invention relates to the use of these receptors to identify compounds in coffee and other bitter foods and beverages that induce a bitter taste. The invention also encompasses test methods that include a further step that evaluates the effect of the identified modulation compounds in humans or other taste tests, and evaluates the effect of the identified compounds on the bitter taste. Also, the invention encompasses the use of the identified compounds in foods, beverages and medicines as flavor or flavor modulators, i.e. to inhibit the bitter taste, for example, the bitter taste associated with coffee beverages and coffee flavor foods. . OBJECTIVES OF THE INVENTION It is an object of the invention to identify compounds that block the activation of hT2R8, hT2R14, or hT2R54 or fragments, variants, orthologs, or chimeras thereof by chlorogenic lactone molecules found in coffee, or a structurally related compound to such chlorogenic lactones that activate at least one of these T2R receptors. It is a specific objective of the invention to identify compounds that block the activation of hT2R58, hT2R14 and hT2R54 or fragments, variants, orthologs, or chimera thereof by 3CoQAL, a chlorogenic lactone comprised in coffee, or a compound structurally related thereto. that activates one or all of these hT2Rs. It is another specific objective of the invention to identify compounds that block the activation of hT2R8, hT2R14, and hT2R54, by 3CQAL and 4CQAL, chlorogenic lactones found in coffee, by or a structurally related compound that specifically activates at least one of these receptors. It is another specific objective of the invention to identify compounds that block the activation of hT2R8, and hT2R54 by 4FQAL, a chlorogenic lactone found in coffee beverages, or a compound structurally related thereto that specifically activates this receptor. It is another specific objective of the invention to use cells or cell membranes comprising or expressing (either stably or transiently) hT2R8, hT2R14 or hT2R54 or a fragment, variant, ortholog, mutant or chimera thereof in assays to identify compounds that block the activation of at least one receptor by chlorogenic lactones and related bitter compounds, for example, 3COQAL, 3CQAL and 4 FQAL. It is still a more specific objective of the invention to use cells, preferably mammary, amphibian or insect cells, for example, HEK293T cells that express a G protein that is coupled to them., for example, Ga15, Ga16, gustducin, transducin or a chimera thereof, for example, g16 gustducin chimera or Gu16 transducin chimera, in cell-based assays that detect changes in the order of intracellular calcium to detect compounds that modulate, preferably they block or inhibit the activation of one of the human taste receptors mentioned in the foregoing by chlorogenic lactones and related compounds comprised in coffee beverages, for example, 3CoQAL, 3CQAL, 4CQAL and 4FQAL. It is another object of the invention to confirm that the identified compounds modulate, preferably inhibit or block, the bitter taste, for example that is induced by the chlorogenic lactones and related bitter compounds, for example, those found in coffee beverages and flavors of test, preferably human test flavors. It is another object of the invention to use compounds identified in the assays described herein, additives or flavor modulators in compositions to inhibit or block the bitter taste induced by the compounds that specifically activate these taste receptors. A preferred object of the invention is to use a compound that inhibits the activation of at least one of the human T2R receptors identified in the above in order to block the bitter taste of the chlorogenic lactones contained in foods, beverages and medicinal substances, preferably coffee or coffee flavor drinks and food.

DETAILED DESCRIPTION OF THE FIGURES Figure 1 contains the structures of various chlorogenic lactones. Figure 2 shows dose responses of hT2R8, hT2R14 and hT2R54 to one of the chlorogenic lactones (3CoQAL).

Figure 3 contains the results of the calcium imaging experiments showing that hT2R8, hT2R14, and hT2R54 respond specifically to two of the chlorogenic lactones, 3CQAL and 4CQAL, depicted in Figure 1. Figure 4 contains the results of an experiment of calcium imaging which resists that hT2R8 and hT2R54 respond to another chlorogenic lactone 4FQAL depicted in Figure 1 in a dose-dependent manner. DETAILED DESCRIPTION OF THE INVENTION Before describing the invention specifically, the following definitions are provided. The term "T2R" family includes polymorphic variants, alleles, mutants, homologs that: (1) have approximately 30-40% amino acid sequence identity, more specifically approximately 40, 50, 60, 70, 75, 80, 85, 90, 95, 96, 97, 98, or 99% amino acid sequence identity to T2Rs disclosed infra, and in the applications of Zuker (Id) (2001) and Adler ( Id.) (2001) incorporated, by reference herein on a window of approximately 25 amino acids, optionally 50-100 amino acids; (2) specifically binding to raised antibodies against an immunogen comprising an amino acid sequence selected from the group consisting of T2R sequences disclosed infra, and conservatively modified variants thereof; (3) specifically hybridize (with a size of at least about 100, optionally at least about 500-1000 nucleotides) under severe hybridization conditions to a sequence selected from the group consisting of the T2R DNA sequences disclosed infra, and variants conservatively modified thereof; (4) comprising a sequence of at least about 40% identical to an amino acid sequence selected from the group consisting of T2R amino acid sequences disclosed infra or (5) conservatives are amplified which hybridize specifically under conditions of severe hybridization to the sequences T2R described. In particular, these "T2R 's" include flavor receptor GPCRs referred to as hT2R8, hTlR14, hT2R54 having the nucleic acid sequences and amino acid sequences provided in this application, and variants, alleles, mutants, orthologs and chimeras thereof which bind specifically to bitter ligands, ie, chlorogenic lactones and derivatives thereof, for example, 3CQAL, 4CQAL, 3CoQAL and 4 FQAL contributing at least to the bitter taste of coffee beverages. While the T2R genes exhibit substantial sequence divergence in both the protein and DNA levels, all of the T2Rs isolated to date have been found to contain certain consensus sequences in particular regions that are identical or have at least 70-75 % sequence identity to the consensual sequence T2R previously identified in Adler et al. (WO 01/77676 Al (2001) and Zuker et al WO 01/18050 A2, both incorporated by reference in their entirety here.) Topologically, certain GPCRs chemosensory cells have an "N-terminal domain", "extracellular domains", a "transmembrane domain" comprising seven transmembrane regions, and corresponding to cytoplasmic and extracellular circuits, "cytoplasmic regions" and a "C-terminal region" ( see, for example, Hoon et al., Cell, 96: 541-51 (1999); Buck &Axel, Cell, 65: 175-87 (1991).) These regions can be structurally identified using all known to those of skill in the art, such as sequence analysis programs that identify hydrophobic and hydrophilic domains (see, for example, Stryer, Biochemistry, (3rd ed. 1988); also see any of a number of Internet-based sequence analysis programs). These regions are useful for being chimeric proteins and for m vitro assays of the invention, for example, ligand binding assays. "Extracellular domains" therefore refers to the T2R polypeptide domains that protrude from the cell membrane and are exposed to the extracellular face of the cell. Such regions would include the "N-terminal domain" that is exposed to the extracellular face of the cell, as well as the extracellular circuits of the transmembrane domain that are exposed to the extracellular face of the cell, i.e., the extracellular circuits between the regions of the cell. transmembrane 2 and 3, the transmembrane regions 4 and 5, and the transmembrane regions 6 and 7. The "N-terminal domain" begins at the N-terminus and extends to a region near the beginning of the transmembrane region. These extracellular regions are useful for in vitro ligand binding assays, both in soluble and solid phases. In addition, the transmembrane regions, described below, can also be involved in ligand binding, either in combination with the extracellular region or alone, and are therefore also useful for in vitro ligand binding assays. "Transmembrane domain", which comprises the seven transmembrane "regions" refers to the domain of T2R polypeptides that are placed within the plasma membrane, and may also include the corresponding cytoplasmic (intracellular) and extracellular circuits, also referred to as "regions". "Transmembrane. The seven transmembrane regions and extracellular and cytoplasmic circuits can be identified using standard methods, as described in Kyte & amp; amp; amp;; Doolittle, J. Mol. Biol, 157: 105-32 (1982)), or in Stryer, supra. "Cytoplasmic domains" refers to the domains of the T2R proteins that face the cell side, for example, the "C-terminal domain" and the intracellular circuits of the transmembrane domain, for example, intracellular circuits between regions 1 and 2, transmembrane, regions 3 and 4, transmembrane and the transmembrane regions 5 and 6. "C-terminal domino" refers to the region extending from the end of the last transmembrane region to the C-terminus of the protein and that is normally located within the cytoplasm. The term "7-transmembrane receptor" means a polypeptide that belongs to a superfamily of transmembrane proteins that have seven regions that extend along the plasma membrane seven times, (thus, the seven regions are called domains of " transmembrane "or" TM "TM I to TM VII). The families of certain taste and olfactory receptors each belong to this superfamily. The 7-transmembrane receptor polypeptides have similar structures and of primary, secondary and tertiary characteristics, as discussed in further detail below. The term "ligand binding region" refers to sequences derived from a chemosensory or taste receptor that substantially incorporates transmembrane domains II through VII (TM II to VII). The region may be capable of binding a ligand, and more particularly, a flavor-inducing compound. The term "plasma membrane translocation domain" or simply "translocation domain" means a polypeptide domain that is functionally equivalent to an exemplary translocalization domain (5 '-MNGTEGPNFYVPFSNKTGVV; SEQ ID NO: 1). These peptide domains, when incorporated into the amino terminus of a polypeptide coding sequence, can more efficiently "accompany" or "translocate" the hybrid protein ("fusion") to the cell plasma membrane. This particular "translocation domain" was derived from the amino terminus of the human rhodopsin receptor polypeptide, a 7-transmembrane receptor. Another translocation domain has been derived from the bovine rhodopsin sequence and is also useful to facilitate translocation. The rhodopsin-derived sequences are particularly efficient in translocating the 7-transmembrane fusion proteins to the plasma membrane. "Functional Equivalence" means that the ability and efficiency of the domain in the translocation of proteins recently translated to the plasma membrane as efficient as SEQ ID NO: l exemplary under similar conditions; Relative efficiencies can be measured (in quantitative terms) and compared, as described herein. Domains that fall within the scope of the invention can be determined by routine classification for their efficiency and translocation of newly synthesized polypeptides to the plasma membrane in a cell (mammal, Xenopus, and the like) with the same efficiency as the twenty amino acid long translocation domain SEQ ID NO: 1. The phrase "functional effects" in the context of assays for testing compounds or modulating the taste transduction mediated by the member of the T2R family includes the determination of any parameter that is indirect or directly under the influence of the receiver, for example, functional, physical and chemical effects. This includes ligand binding, changes in ion flux, membrane potential, current flow, transcription, G protein binding, GPCR phosphorylation or dephosphorylation, signal transduction, receptor-ligand interactions, second messenger concentrations (by example, cAMP, cGMP, IP3, or intracellular Ca2 +), in vitro, in vivo, and ex vivo and also includes other physiological effects such as increases or decreases of the neurotransmitter or release of hormone. By "determining the functional effect" a test is proposed for a compound that increases or decreases a parameter that is indirect or directly under the influence of a member of the T2R family, for example, functional, physical and chemical effects. Such functional effects can be measured by any means known to those skilled in the art, for example, changes in spectroscopic characteristics (e.g., fluorescence, absorbency, refractive index), hydrodynamics (e.g., shape), chromatographic, or properties of solubility, patch fixation, voltage-sensitive dyes, whole cell currents, radioisotope output, inducible markers, oocyte T2R expression: T2R cell expression of tissue culture; Trans-transcriptional activation of T2R genes; ligand binding assays; voltage, membrane potential and conductance changes; ion flow assays; changes in the second cell-like messengers such as cAMP, cGMP, and mositol triphosphate (IP3); changes in intracellular calcium levels; neurotransmitter release and the like. "Inhibitors", "activators" and "modulators" of T2R protein receptors are used interchangeably to refer to inhibit, or activate or modulate, molecules identified using in vitro or om vivo assays for flavor transduction, eg, ligands, agonists, antagonists. , and their counterparts and imitations. Inhibitors are compounds that, for example, bind to, partially or totally block stimulation, decrease, prevent, retard activation, inactivate, desensitize, or down-regulate taste transduction, for example, antagonists. Activators are compounds that, for example, bind to, stimulate, increase,. open, facilitate, increase activation, sensitize, or up-regulate flavor transduction, for example, agonists. Modulators include compounds that, for example, alter the interaction of a receptor with extracellular proteins bind activators or inhibitor; (for example, ebnerin and other members of the hydrophobic carrier family); G proteins; kinases (eg, rhodopsin kinase homologs and beta adrenergic beta receptor kinases that are involved in the deactivation and sensitization of a receptor); and arrestins that also deactivate and desensitize the receptors. Modulators include genetically modified versions of members of the T2R family, for example, with altered activity, as well as naturally occurring ligands and synthetic antagonists, agonists, small chemical molecules and the like. Such assays for inhibitors and activators include, for example, expressing members of the T2R family in cells or cell membranes, applying putative modulator compounds in the presence or absence of compounds that modulate, for example, bitter compounds, and then determining the functional effects on flavor transduction, as described in the above. Samples or assays comprising members of the T2R family that are treated with a potential act, inhibitor, or modulator are compared to the control samples without the inhibitor, activator, modulator to examine the degree of modulation. Control samples (not treated with modulators) are assigned a relative T2R activity value of 100%. Inhibition of a T2R is achieved when the value of the T2R activity relative to the control is about 80%, optionally 50% or 25-0%. Activation of a T2R is achieved when the value of the T2R activity relative to the control is 110%, optionally 150%, optionally 200-500%, or 1000-3000% higher. The terms "purified" "substantially purified" and "isolated" as used herein refers to the state that is free from other, similar compounds with the compound of the invention are normally associated in their natural state. Preferably, "purified" "substantially purified" and "isolated" means that the composition comprises at least 0.5%, 1%, 5%, 10%, or 20%, and much more preferably at least 50% or 75% of the mass, by weight of a given sample. In a preferred embodiment, these terms refer to 1 compound of the invention comprising at least 95% of the mass, by weight, of a given sample. As used herein, the terms "purified", "substantially purified," and "isolated", when referring to a nucleic acid or protein, of nucleic acids or proteins, and refers to a different purification state or concentration than that which occurs naturally in the mammalian body, especially human. Any degree of purification or concentration greater than that which occurs naturally in the body of mammals, especially to human, which includes (1) the purification of other structures of associated compounds or (2) the association with structures of compounds to which they are not normally associated in the mammalian body, especially human, are within the meaning of " isolated. " The nucleic acid or protein or classes of nucleic acids or proteins, described herein, may be isolated, otherwise associated with structures or compounds to which they are not normally associated in nature, according to a variety and known methods and processes. by those of skill in the art. As used herein, the term "isolated", when referring to a nucleic acid or polypeptides, refers to a state of purification or concentration different from that which occurs naturally in the mammalian body, especially human. Any grade of purification or concentration is greater than that occurring naturally in the body, including (1) the purification of other structures or associated compounds that occur naturally, or (2) the association with structures or compounds to which they are not normally associated in the body they are within the meaning "isolated" as used in the present. The nucleic acids or polypeptides described herein may be isolated or otherwise associated with structures or compounds to which they are not normally associated in nature. According to a variety of methods and processes known to those of skill in the art. As used herein, the terms "amplify" and "amplification" refers to the use of any suitable amplification methodology to generate or detect nucleic acid, recombinant or naturally expressed, as described in detail, below. For example, the invention provides methods and reagents (eg, pairs of specific oligonucleotide primers) to amplify (e.g., by polymerase chain reaction, PCR) naturally expressed (e.g., genomic or mRNA) or recombinant nucleic acids. (e.g., cDNA) of the invention (e.g., linker sequences of flavor-inducing compound of the invention) in vivo or in vitro. The term "expression vector" refers to any recombinant expression system for the purpose of expressing a nucleic acid sequences of the invention in vitro on vivo, constitutively or inductibly, in any cell, including prokaryotic, yeast cell, fungal, plant, insect or mammal. The term includes linear or circular expression systems. The term includes expression systems that remain episomal or integrated into the genome of the host cell. Expression systems may have the ability to self-replicate or not, that is, to drive only transient expression in a cell. The term includes "cassettes" of recombinant expression that contain only the minimum elements necessary for transcription of the recombinant nucleic acid. The term "library" means a preparation which is a mixture of different nucleic acid or polypeptide molecules, such as the library of enLace regions of receptor ligands particularly taste, sensory generated recombinants generated by the application of nucleic acid with primer pairs generated, or an isolated collection of vectors that incorporate the amplified ligand binding regions or a mixture of cells each randomly transfected with at least one vector encoding a flavor receptor. The term "nucleic acid" or "nucleic acid sequences" refers to a deoxy ribonucleotide ribonucleotide, oligonucleotide in the form of either one or two strands. The term includes nucleic acids, i.e., oligonucleotides, which contain known analogs of natural nucleotides. The term also includes structures similar to nucleic acids with synthetic backbones. Unless otherwise indicated, a particular nucleic acid sequence also implicitly includes vanishing ones conservatively modified from themselves (eg, degenerate codon substitutions) and complementary sequences, as well as the implicitly indicated sequence. Specifically, generated codon substitutions can be achieved by generating, for example, sequences in which the third position of one or more selected codons is substituted with mixed base residues and / or deoxyinosine (Batzer et al., Nucleic Acid Res. , 19: 5081 (1991), Ohtsuka et al., J. Biol. Chem., 260: 2605-08 (1985), Rossolini et al., Mol. Cell. Probes, 8: 91-98 (1994)). The term nucleic acid is used interchangeably with the gene, cDNA, mRNA, oligonuc Leotide, and polynucleotide. The terms "polypeptide", "peptide" and "protein" are used interchangeably herein to refer to a polymer of amino acid residues. The terms apply to polymers of amino acids in which one or more amino acid residue is an artificial chemical copy of a naturally occurring amino acid, as well as polymers of naturally occurring amino acids and non-naturally occurring amino acid polymer. The "translocation domain", "ligand binding region", and chimeric receptor compositions described herein also include "analogues" or "conservative variants" "mimetics" ("mimetic peptides") with substantially corresponding structures and activity. to the exemplary sequences. Thus, the terms "conservative variants" or "analogue" or "mimetic" refers to a polypeptide having a modified amino acid sequence, such that the change (s) does not substantially alter the structure of the polypeptide (of the variant conservative) and / or activity, as defined herein. These include conservatively modified variations of an amino acid sequence, i.e., amino acid substitutions, additions or deletions of those residues that are not critical for protein activity, or substitution of amino acids with residues having similar properties (eg, acidic, basic, positively or negatively charged, polar or non-polar, etc.) such that substitutions of even critical amino acids do not substantially alter the structure and / or activity. More particularly, "conservatively modified variants" apply to both amino acid and nucleic acid sequences. With respect to particular nucleic acid sequences, conservatively modified variants refer to those nucleic acids that encode identical or essentially identical amino acid sequences, or where the nucleic acid does not encode an amino acid sequence, the essentially identical sequences. Due to the degeneracy of the genetic code, a large number of functionally identical nucleic acids encode any given protein. For example, the GCA, GCC, GCG and GCU codons all encode the amino acid alanine. Thus, in each position where an alanine is specified by a codon, the codon can be altered to any of the corresponding codons described without altering the encoded polypeptide. Such variations of nucleic acid are "silent variations", which are not species of conservatively modified variations. Each nucleic acid sequence herein that encodes a polypeptide also describes each possible silent variation of the nucleic acid. One of skill will recognize that each codon in a nucleic acid (except AUG, which is ordinarily the only codon for methionine, and TGG, which is ordinarily the only codon for tryptophan) can be modified to produce a functionally identical molecule. Accordingly, each silent variation of a nucleic acid encoding a peptide is implicit in each described sequence. Conservative substitution tables that provide functionally similar amino acids are well known in the art. For example, an exemplary guide for selecting conservative substitutions includes (original residue followed by exemplary substitution): wing / gly or be; arg / lys; asn / gln or his; asp / glu; cys / ser; gln / asn; gly / asp; gly / ala or pro; his / asn or gin; ile / leu or val; leu / ile or val; lys / arg or gin or glu; met / leu or tyr or lie; phe / met or leu or tyr; be / thr; thr / be; trp / tyr; tyr / trp or phe; val / ile or leu. An alternative exemplary guide uses the following six groups, each containing amino acids that are conservative substitutions for each other: 1) Alanine (A), Serine (S), Threonine (T); 2) Aspartic acid (D), Glutamic acid (E); 3) Asparagine (N), Glutamine (Q); 4) Arginine (R), Lysine (I); ) Isoleucine (I), Leucine (L), Methionine (M), Valine (V); Y 6) Phenylalanine (F), Tyrosine (Y), Tpptofano (W); (see also, for example, Creighton, Proteins, W. H. Freeman and Company (1984), Schultz and Schimer, Principies of Protein Structure, Springer-Verlag (1979)). One of skill in the art will appreciate that the substitutions identified in the foregoing are not only possible conservative substitutions. For example, for some purposes, one can consider all amino acids loaded as conservative substitutions for each if they are positive or negative. In addition, individual substitutions, deletions, additions that alter, add or delete a single amino acid or a small percentage of amino acids in a modified sequence can also be considered "conservatively modified variations". The terms "mimetic" and "mimetic peptide" refer to a synthetic chemical compound having substantially the same structural and / or functional characteristics of the polypeptides, for example, translocation domains, ligand binding regions, or chimeric receptors of the invention. The mimetic may be either composed entirely of non-natural, synthetic analogs of amino acids, or may be a chimeric molecule of partially natural peptide amino acids and partially unnatural analogs of amino acids. The mimetic may also incorporate any number of conservative substitutions of natural amino acids as long as such substitutions also do not substantially alter the structure and / or mimetic activity. As with the polypeptides of the invention which are conservative variants, routine experimentation will determine whether a mimetic is within the scope of the invention, that is, that its structure and / or function is not substantially altered. The polypeptide mimetic compositions may contain any combination of non-natural structural components, which are typically of three structural groups: a) residue linking groups other than the natural amide linking bonds ("peptide bond"); b) non-natural residues instead of naturally occurring amino acid residues; or c) residues that induce secondary structural mimicry, that is, to induce or stabilize a secondary structure, eg, a beta-turn, gamma spin, beta-sheet alpha-helix conformation, and the like. A polypeptide can be characterized as a mimic when all or some of its residues are bound by chemical means other than the natural peptide bond. The individual peptide mimetic residues can be linked by peptide bonds, other chemical bonds or coupling means, such as, for example, glutaraldehyde, N-hydroxysuccinimide esters, bifunctional maleimides, N, N '-dicyclohexylcarbodnmide (DCC) or N, N'-diisopropylcarbodnmide (DIC). Linking groups that can be an alternative to traditional amide linkage linkages ("peptide link") include, for example, ketomethylene (eg, --C (.bdbd.O) --CH2for-C ( dbd.O) --NH--), ammonomethyl (CH2NH), ethylene, olefin (CH.dbd.CH), ether (CH20), thioether (CH2-S), tetrazole (CN4), tlazole, retroamide, thioamide , or ester (see, for example, Spatola, Chemistry and Biochemistry of Amino Acids, Peptides and Proteins, Vol. 7, 267-357, Marcell Dekker, Peptide Backbone Modifications, NY (1983)). A polypeptide can also be characterized as a mimetic by containing all or some unnatural residues at the site of naturally occurring amino acid residues; Unnatural residues are well described in scientific and patent literature. A "label" or a "detectable portion" is a composition detectable by spectroscopic, photochemical, biochemical, immunochemical, or chemical means. For example, useful labels include 32P, fluorescent dyes, dense electron reagents, enzymes (eg, as commonly used in ELISA), biotin, digoxigenin, or haptens and proteins that can be made detectable, for example, by incorporating a radiolabel into the peptide used to detect antibodies specifically reactive with the peptide. A "labeled nucleic acid probe or oligonucleotide" is one that is linked, either covalently through a linker or a chemical or non-covalently linked, through ionic bonds, van der Waals, electrostatic, or hydrogen to a brand such that the presence of the probe can be detected by detecting the presence of the label linked to the probe. As used herein a "nucleic acid probe or oligonucleotides" is defined as a nucleic acid capable of binding to a target nucleic acid of complementary sequence through one or more types of chemical bonds, usually through base pairing complementary, usually through the formation of hydrogen bonding. As used herein, a probe can include natural (ie, A, G, C, or T) or modified bases (7-deazaguanosine, inosine, etc.). In addition, bases in a probe can be linked by a different link to a phosphodiester linkage, so long as it does not interfere with hybridization. Thus, for example, the probes may be peptide nucleic acids in which the constituent bases are linked by peptide bonds other than the phosphodiester linkages. It will be understood by one of skill in the art that the probes can link target sequences lacking complete complementarity with the probe sequence depending on the severity of the hybridization conditions. The probes are optionally labeled directly as isotopes, chromophores, lumiphores, chromogens, or are labeled indirectly such as with biotin to which a streptavidin complex can then bind. When analyzing for the presence or absence of the probe one can detect the presence or absence of the selected sequence or sub-sequence. The term "heterologous" when used with reference to portions of a nucleic acid indicates that the nucleic acid comprises two or more subsequences that are not in the same relationship to one another in nature. For example, the nucleic acid is typically produced recombinantly, having two or more sequences of unrelated genes arranged to make a new nucleic acid work, for example a promoter from one source and a coding reaction from another source. Similarly, a heterologous protein indicates that the protein comprises two or more subsequences that are not in the same relationship to one another in nature (e.g., a fusion protein). A "promoter" is defined as an array of nucleic acid sequences that direct the transcription of a nucleic acid. As used herein, a promoter includes necessary nucleic acid sequences near the site of transcription initiation. As in the case of a polymerase type 11 promoter, a TATA element. A promoter also optionally includes distal enhancer or repressor elements, which can be localized as many as several thousand base pairs from the start site of transcription. A "constitutive" promoter is a promoter that is active under most environmental and developmental conditions. A "promoter" promoter is a promoter that is active under environmental or developmental regulation. The term "operably linked" refers to a functional link between a nucleic acid expression control sequence (such as a promoter, or array of transcription factor binding sites) and a second nucleic acid sequence, wherein the The expression control sequence directs the transcription of the nucleic acid corresponding to the second sequence. As used herein, "recombinant" refers to a polynucleotide synthesized or otherwise manipulated in vitro (eg, "recombinant polynucleotide") to methods for using recombinant polypeptides to produce ten products in cells or other biological systems, a polypeptide ("recombinant protein") encoded by a recombinant polypeptide "recombinant medium" also includes ligation of nucleic acids having vain regions or coding domains or promoter sequences from different sources in an expression cassette or vector for expression of, for example, inducible or constitutive expression of a fusion protein comprising a translocation domain of the invention and an amplified nucleic acid sequence using a primer of the invention. The phrase "selectively (or specifically) a hybrid" refers to the binding, duplication or hybridization having molecule only to a particular nucleotide sequence under severe hybridization conditions when that sequence is present in a complex mixture (e.g. Total cellular RNA or library). The phrase "Severe hybridization conditions" refers to the conditions under which a zonda will hybridize to objective subsequence, typically in a complex mixture of nucleic acid, but not to other sequences. The severe conditions are the dependent sequence and will be different in different circumstances. The longer sequences are specifically hybridized at higher temperatures. An extensive guide to nucleic acid hybridization is found in Tijssen, Techniques in Biochemistry and Molecular Biology-Hybridization ith Nucleic Probes, "Overview of Principles of Hybridization and the Strategy of Nucleic Acids Assays" (1993). Generally, severe conditions are selected to make approximately 5-10 ° C, lower than the thermal melting point (Tm) for the specific sequence at a defined pH ionic strength. The Tm is the temperature (low ionic strength defined pH, and nucleic concentration) in which 50% of the probes complementary to the target are hybridized to the objective sequence in the equilibrium (as the target sequences are present in excess, in Tm, 50 % of the probes are occupied in the balance). The severe conditions will be those in which the salt concentration is less than about 1.0 M sodium ion, typically approximately 0.01 to 1.0 M concentration of sodium ion (or other salts) at a pH of 7.0 to 8.3 and the temperature is at least approximately 30 ° C. For short probes and (for example, 10 to 50 nucleotides) and at least approximately 6 ° C for long probes (for example, greater than 50 nucleotides). Severe conditions can also be achieved with the addition of destabilizing agents such as formamide. For selective or specific hybridization, a positive signal is at least two times background, optionally 10 times the background hybridization. Exemplary severe hybridization conditions may be as follows: 50% formamide, 5xSSC, and 1% SDS, incubate at 42 ° C, or, 5xSSC, 1% SDS, incubate at 65 ° C, with wash in 0.2xSSC, and 0.1% SDS at 65 ° C. Such hybridizations and washing steps can be carried out by, for example, 1, 2, 5, 10, 15, 30, 60; or more minutes. Nucleic acids that do not hybridize to each other under severe conditions are still substantially related if the polypeptides they code are substantially related. This occurs, for example, when a copy of a nucleic acid is created using the maximum codon degeneracy allowed by the genetic code. In such cases, nucleic acids are typically hybridized under moderately severe hybridization conditions, exemplary "moderately severe hybridization conditions" include hybridization in a 40% formamide buffer solution, 1 M NaCl, 1% SDS at 37 ° C. , and a wash in IxSSC at 45 ° C. Such hybridizations and washing steps can be carried out by, for example, 1, 2, 5, 10, 15, 30, 60, or more minutes. A positive hybridization is at least twice background. Those of ordinary skill will readily recognize that alternative hybridization and washing conditions can be used to provide conditions of similar severity. "Antibody" refers to a polypeptide comprising a framework region of an immunoglobulin gene or fragments thereof that specifically binds and recognizes an antigen. The recognized immunoglobulin genes include kappa, lambda, alpha, gamma, delta, epsilon, and mu constant region genes, as well as variable range immunoglobulin genes of wide variety. Light chains are classified as either kappa or lambda. The heavy chains are classified as gamma, mu, alpha, delta, or epsilon, which in turn define the classes of immunoglobulin, IgG, IgM, IgA, IgD and IgE, respectively. An exemplary immunoglobulin structural unit (antibody) comprises a tetramer. Each tetramer is composed of two identical pairs of polypeptide chains, each pair having a "light" chain (approximately 25 kDa) and a "heavy" chain (approximately 50-70 kDa). The N-terminal of each chain defines a variable region of approximately 100 to 110 or more amino acids mainly responsible for the recognition of the antigen. The terms variable light chain (VL) and variable heavy chain (VH) refer to these light and heavy chains respectively. A "chimeric antibody" is an antibody molecule in which (a) the constant region, or a portion thereof, is altered, replaced or exchanged so that the antigen binding site (variable region) is linked to a constant region of a different or altered class; effector function and / or species, or a completely different molecule that adds new properties to the chimeric antibody, for example, an enzyme, toxin, hormone, growth factor, drug, etc; or (b) the variable region or a portion thereof, is altered, replaced or exchanged with a variable region having a different or altered antigen specificity. An "anti? -T2R" antibody is an antibody or antibody fragment that specifically binds a polypeptide encoded by a T2R gene, cDNA, or a sequence thereof. The term "immunoassay" is an assay that uses an antibody to specifically bind an antigen. The immunoassay is characterized by the use of specific binding properties of a particular antibody to isolate, direct, and / or quantify the antigen. The phrase "specifically binds (or selectively) an antibody" or, "specifically (or selectively) immunoreactive with", when referring to a protein or peptide, refers to a binding region that is determinative of the presence of the protein in a heterogeneous population of proteins and other biologics. Thus, under designated immunoassay conditions, the specified antibodies bind to a particular protein at least twice the background and do not substantially bind in a significant amount to other proteins present in the sample. Specific links to an antibody ba or such conditions may require an antibody that is selected for its specificity for a particular protein. For example, polyclonal antibodies raised to a T2R family member of specific species such as rat, mouse, or human can be selected to obtain only those polyclonal antibodies that are specifically immunoreactive with the T2R polypeptide or an immunogenic portion thereof and not with other proteins, except for photologists or polymorphic variants and alleles of the T2R polypeptide. This selection can be achieved by subtracting antibodies that cross-react with the T2R molecules of other species or other T2R molecules. Antibodies can also be selected that recognize only members of the T2R GPCR family but not GPCRs of other families. A variety of immunoassay formats can be used to select antibodies especially immunoreactive with a particular protein. For example, solid-phase ELISA immunoassays are routinely used to select antibodies specifically immunoreactive with a protein (see, for example, Harlow & amp; amp;; Lane, Antibodies, A Laboratory Manual, (1988), for a description of immunoassay formats and conditions that can be used to determine the specific immunoreactivity). Typically a specific or selective reaction will be at least twice the background of the signal or noise and more typically more than 10 to 100 times of background. The phrase "selectively associated with" refers to the ability of a nucleic acid to "selectively hybridize" with another as defined above, or the ability of an antibody to selectively (or specifically) bind to a protein, as defined in the above. The term "expression vector" refers to any recombinant expression system for the purpose of expressing a nucleic acid sequence of the invention either in vitro or in vivo, constitutively or physically, in any cell including prokaryotic cell, yeast, fungal, of plant, insect or mammal. The term includes linear or circular expression systems. The term includes expression systems that remain episomal or integrated into the genome of the host cell. Expression systems may have the ability to self-replicate or not, that is, to drive only transient expression in a cell. The term includes recombinant expression cassettes that contain only the minimum elements necessary for the transcription of recombinant nucleic acid. By "host cell" a cell is proposed that contains an expression vector and supports the replication or expression of the expression vector. Host cells can be prokaryotic cells such as E. coli, or eukaryotic cells such as yeast, insect, amphibian, mammalian cells such as CHO, HeLa, HEK-293, and the like, eg, cultured cells, explants and in vivo cells. Based on the above, the present invention provides assays for identifying compounds that modulate, preferably block, the specific activation of the human bitter taste receptor previously identified by bitter compounds for example, chlorogenic lactones found in coffees and other foods and beverages and compounds structurally related. Particularly, the invention provides cell-based assays for identifying compounds that modulate the activation of specific human T2Rs by compounds found in coffee, ie, chlorogenic lactones such as are contained in Figure 1. In particular, the invention involves the discovery of that the activation of hT2R8, hT2R14 or hT2R54 is effected by 3CoQAL chlorogenic lactone or structurally related compounds; hT2R8, hT2R14 or hT2R54 by the chlorogenic lactones 3CQAL and 4FQAL or structurally related compounds; or hT2R8 and hT2R54 to the chlorogenic lactone 4FQAL or structurally related compounds. As noted in the structure of these specific chlorogenic lactones and others are contained in Figure 1. It is anticipated that the identified compounds using assays according to the invention will modulate the bitter taste associated with these taste receptors in human subjects. This will be confirmed in taste tests. That the above taste receptors respond specifically to the bitter taste ligands, ie the chlorogenic lactones found in coffee such as 3CQAL, 4CQAL, 3CoQAL and 4 FQAL was determined using the HEK293 expression system and the imaging methods of calcium reported in other publications as well as the patent applications filed by the present assignee, that is, US serial no. 10 / 191,058 and 09 / 825,882, both incorporated by reference in their totals herein. More particularly, the present inventors transfected HEK293 cells with a particular hT2R in conjunction with a chimeric G protein (G16gust44) comprising the Ga16 protein sequence modified by the replacement of the 44 amino acid residues of carboxy with those of gustucin, and responses of these cells to specific bitter ligands were recorded by calcium imaging methods. As shown in Figure 2, it was found that hT2R8, hT2R14, and hT2R54 respond to 3CoQAL in different concentrations and in a dose-dependent manner. By contrast, these cells do not respond to sucrose in the same concentration. Therefore, these cells or other cells that functionally express these receptors can be used in assays to identify compounds that modulate the activation of these receptors by the bitter compounds, ie chlorogenic lactones, for example, which block the activation of 3CoQAL from these receptors. Also, as shown in Figure 3, it was observed that HEK-293 cells expressing hT2R8 hT2R14 or hT2R54 respond specifically to chlorogenic lactones 3CQAL and 4CQAL in 2mM concentration. By contrast, these cells do not respond to sucrose (control). Furthermore, as shown in Figure 4, it was observed that they utilize these same calcium imaging methods that the HEK-293 cells expressing hT2R8 or hT2R54 responded specifically to the chlorogenic lactone 4FQAL in a dose-dependent manner. By contrast, these cells do not respond to sucrose. These results indicate that cells functionally expressing either of the hT2R8, hT2R14, and hT2R54 flavor receptors can be used in assays to identify ligands that modulate hT2R8, hT2R14, or hT2R54 associated with bitter taste, eg, which are induced by the chlorogenic lactones or structurally related compounds found in coffee beverages and other chlorogenic lactone that contains bitter taste in beverages, foods or medicinal substances. Preferably, these assays will utilize a test cell that expresses a DNA encoding an hT2R having one of the amino acid sequences identified infra. However, it is anticipated that the fragments, orthologs, variants or chimeras of these receptor polypeptides that retain the functional properties of these sweet-umami or umami-sweet flavor receptors, ie, respond to some sweet or umami compounds or The same compounds will also be useful in these tests. Examples of such variants include splice variants, single nucleotide polymorphisms, allelic variants, and mutations produced by recombinant or chemical means, or occurring naturally. The means for the isolation and expression of T2Rs, which are used in the assays of the present invention and assays that are contemplated for use in the present invention to identify compounds that inhibit the activation of these receptors, are discussed below. Isolation and Expression of the T2Rs The isolation and expression of the T2Rs, or fragments or variants thereof, of the invention can be performed by well-established cloning procedures using probes or primers constructed based on the T2R nucleic acid sequences disclosed in application. Related T2R sequences can also be identified from genomic databases of humans or other species using the sequences disclosed herein and known computer-based search technologies, for example, BLAST sequence search. In a particular embodiment, the pseudogenes disclosed herein, can be used to identify functional alleles or related genes. The expression vectors can then be used to infect or transfect host cells for the functional expression of these sequences. These genes and vectors can be made and expressed in vitro or in vivo. One of skill will recognize that the desired phenotypes to alter and control nucleic acid expression can be obtained by modulating the expression or activity of genes and nucleic acids (e.g., promoters, enhancers and the like) within the vectors of the invention. Any of the known methods described to increase or decrease the expression or activity can be used. The invention can be practiced in conjunction with any method or protocol known in the art, which is well described in the scientific and patent literature. Alternatively, these nucleic acids can be synthesized in vitro by well known chemical synthesis techniques, as described in, for example, Carruthers, Cold Spring Harbor Symp. Quant. Biol. 47: 411-18 (1982); Adams, Am. Chem. Soc, 105: 661 (1983); Belousov, Nucleic Acids Res. 25: 3440-3444 (1997); Frenkel, Free Radie. Biol. Med. 19: 373-380 (1995); Blommers, Biochemistry 33: 7886-7896 (1994); Narang, Meth. Enzymol. 68:90 (1979); Brown, Meth. Enzymol. 68: 109 (1979); Beaucage, Tetra. Lett. 22: 1859 (1981); U.S. Patent No. 4,458,066. Two-stranded DNA fragments can then be obtained either by synthesizing the complementary strand and annealing the strands together under appropriate conditions, or by adding the complementary strand using DNA polymerase with an appropriate primer sequence. Techniques for the manipulation of nucleic acids, such as, for example, for sequence generation mutations, subcloning, labeling probes, sequencing, hybridization and the like are well described in the scientific and patent literature, see, for example, Sambrook, ed. , Molecular Cloning: A Laboratory Manual (2nd ed.), Vols. 1-3, Cold Spring Harbor Laboratory (1989); Ausubel, ed., Current Protocols m Molecular Biology, John Wiley & Sons, Inc., New York (1997); Tijssen, ed., Laboratory Techniques in Biochemistry and Molecular Biology: Hybridization With Nucleic Acid Probes, Part I, Theory and Nucleic Acid Preparation, Elsevier, N. Y. (1993). The nucleic acids, vectors, capsids, polypeptides, and the like can be analyzed and quantified by any of a number of general means well known to those of skill in the art. These include, for example, analytical biochemical methods such as NMR, spectrophotometry, radiography, electrophoresis, capillary electrophoresis, high performance liquid chromatography (HPLC), thin layer chromatography (TLC), and h: perdifusion chromatography, various immunological methods, for example, fluid or gel precipitation reactions, immunodiffusion, immunoelectrophoresis, radioimmunoassays (RIAs), enzyme-linked immunosorbent assays (ELISAs), immuno-fluorescent assays, Southern analysis, Northern analysis, spot spot analysis, electrophoresis gel (eg, SDS-PAGE), RT-PCR, quantitative PCR, other nucleic or target signal amplification methods, radiolabeling, scintillation counting, and affinity chromatography. Oligonucleotide primers can be used to amplify nucleic acids encoding a T2R ligand binding region. The nucleic acids described herein can also be cloned or quantitatively measured using amplification techniques. Amplification methods are also well known in the art and include, for example, polymerase chain reaction (PCR) (Imns ed., PCR Protocols, a Guide to Methods and Applications, Academic Press, N. Y. (1990); Innis ed., PCR Strategies, Academic Press, Inc., N. Y. (1995)); ligase chain reaction (LCR) (Wu, Genomics, 4: 560 (1989); Landegren, Science, 241: 1077 (1988); Bamnger, Gene, 89: 117 (1990)); transcription amplification (Kwoh, PNAS, 86: 1173 (1989)); self-sustained sequence replication (Guatelli, PNAS, 87: 1874 (1990)); amplification of Q Beta replicase (Smith, J. Clin Microbiol., 35: 1477-91 (1997)); automated Q-beta replication amplification assay (Burg, Mol, Cell, Probes, 10: 257-71 (1996)); and other techniques mediated with RNA polymerase (e.g., NASBA, Cangene, Mississauga, Ontario). See also, Berger, Methods Enzymol., 152: 307-16 (1987); Sambrook; Ausubel; U.S. Patent Nos. 4,683,195 and 4,683,202; Sooknanan, Biotechnology, 13: 563-64 (1995). Once amplified, the nucleic acids, either individually or as libraries, can be cloned according to methods known in the art, if desired, in any of a variety of vectors using routine molecular biological methods; Methods for cloning amplified nucleic acids in vitro are described, for example, in U.S. Patent No. 5,426,039. To facilitate the cloning of the amplified sequences, the restriction enzyme sites can be "built in" the pair of the PCR primer. For example, the Pst I and Bsp El sites were designated in the exemplary primer pairs of the invention, these particular restriction sites have a sequence that, when linked, are "in structure" with respect to the "donor" coding sequence. "7-membrane receptor in which they are separated (the ligand binding region coding sequence is internal to the 7-membrane polypeptide as well, if it is desired that the construct be translated downstream of a restriction enzyme separation site. , outside the results of the structure should be avoided, this may not be necessary if the ligand binding region inserted comprises substantially the majority of the transmembrane region VII). The primers can be designed to retain the original sequence of the 7-membrane "donor" receptor. Alternatively, the primers can be encoded amino acid residues that are conservative substitutions (eg, hydrophobic for the hydrophobic residue, see discussion above) or functionally benign substitutions (eg, do not prevent plasma membrane insertion, cause excision by the peptidase, cause abnormal folding of the receptor, and the like). The primer pairs can be designed to selectively amplify the ligand binding regions of the T2R proteins. These linking regions can be varied for different ligands; thus, which may be a minimal binding region for a ligand, may be very limiting for a second potential ligand. Thus, the linking regions of different sizes comprising different domain structures can be amplified; for example, transmembrane (TM) II to VII, III to VII, III to VI or II to VI domains, or variations thereof (eg, only a subsequence of a particular domain, by mixing the order of the domains, and similar ones), of a T2R of 7-transmembrane. As structures and domain sequences of many 7-membrane T2R proteins are known, the skilled person can easily select the domain flanking and internal domain sequences as model sequences to designate the generated amplification primer pairs. For example, a nucleic acid sequence encoding domain II to VII regions can be generated by PCR amplification using a primer pair. To amplify a transmembrane domain I (TM I) sequence comprising nucleic acid, a degenerate primer can be designed from a nucleic acid encoding the amino acid sequence of the consensus sequence of the T2R 1 family described in the foregoing. Such a degenerate primer can be used to generate a binding region that incorporates TM I into TM III, TM I to TM IV, TM I to TM V, TM I to TM VI or TM I to TM VII). Other degenerate primers can be designed based on the other consensus sequences of the T2R family provided herein. Such a degenerate primer can be used to degenerate a binding region that incorporates TM III to TM IV, TM III to TM V, TM III to TM VI or TM III to TM VII. The paradigms for designing the degenerate primer pairs are well known in the art. For example, a Hybrid Consensus-Degenerate Oligonucleotide Primer (CODEHOP) computer program is accessible as http://blocks.fhcrc.org/codehop.html, and is linked directly from the BlockMaker multiple sequence alignment site. for the prediction of the hybrid primer that begins with a set of related protein sequences, such as the known receptor-receptor ligand binding regions (see, eg, Rose, Nucleic Acids Res., 26: 1628-35 (1998)); Singh, Biotechmques, 24: 318-19 (1998)). Means for synthesizing the primary oligonucleotide primers are well known in the art. Base pairs "natural" or synthetic base pairs can be used. For example, the use of artificial nucleobases offers a versatile procedure for manipulating the primer sequences and generating a more complex mixture of amplification products. Several families of artificial nucleobases are capable of assuming multiple hydrogen enLace orientations through internal link rotations to provide a means for degenerate molecular recognition. The incorporation of these analogs in a single position of a PCR primer allows the generation of a complex library of amplification products. See, for example, Hoops, Nucleic Acids Res., 25: 4866-71 (1997). Non-polar molecules can also be used to mimic the shape of natural DNA bases. An imitation of non-hydrogen bonding form for adenine can efficiently and selectively replicate against a non-polar imitation for thiamine (see, eg, Morales, Nat. Struct. Biol, 5: 950-54 (1998)) ). For example, two degenerate bases can be 6H, pyrimidine base, 8H-3,4-dihydropyrimido [4, 5-c] [1, 2] oxaz? N-7-one or N6-methox? -2, Purine-based 6-diaminopurine (see, eg, Hill, PNAS, 95: 4258-63 (1998)). Exemplary degenerate primers of the invention incorporate the nucleobase analogue 5'-Dimetoxy? Tr? T? LN-benzoyl-2'-deoxy-Citidma, 3 '- [(2-cyanoet? L) - (N, N- dnsopropyl)] -phosphoramidite (the term "P" in the sequences, see above). This pyrimidine analog hydrogen is linked to pupils, which include A and G residues. The polymorphic variants, alleles, interspecies homologs that are substantially identical to a taste receptor disclosed herein can be isolated using the nucleic acid probes described. in the above. Alternatively, expression libraries can be used to clone T2R polypeptides and polymorphic allele variants, and homologues of mterespecies thereof, by detecting homologs expressed mmunologically with purified antisera or antibodies made against a T2R polypeptide, which also recognizes and selectively binds the homologue T2R. The nucleic acids encoding the ligand binding regions of the test receptors can be generated by amplifying (eg, PCR) the appropriate nucleic acid sequences using appropriate primer pairs (perfect or degenerate). The amplified nucleic acid can be genomic DNA from any cell or tissue or mRNA or cDNA derived from the taste receptor expression cells. In one embodiment, hybrid protein coding sequences comprising nucleic acids encoding chimeric or native T2Rs fused to a translocation sequence can be constructed. Hybrid T2Rs are also provided which comprise the translocalization portions and the flavor-binding compound regions of flavor of other chemosensory receptor families, particularly taste receptors. These nucleic acid sequences can be operably linked to transcriptional or transduction control elements, for example, transcription and translation initiation sequences, promoters and enhancers, transcription and translation terminators, polyadenylation sequences and other sequences useful for transcribing DNA in RNA. In the construction of two recombinant expression cassettes, the vectors, and transgenic, a promoter fragment can be used to direct the expression of the desired nucleic acid in all desired cells or tissues. In another embodiment, the fusion proteins can include the C-terminal or N-terminal translocation sequences. In addition, the fusion proteins may comprise additional elements, for example, for protein detection, purification or other applications. Domains that facilitate detection and purification include, for example, metal chelating peptides such as polyhistidine tracts, histidine-tryptophan modules, or other domains that allow purification on immobilized metals.; protein that binds maltose; Protein A domains that allow purification over immobilized immunoglobulin; or the domain used in the FLAGS extension / affinity purification system (Immunex Corp, Seattle Wash.). The inclusion of cleavable linker sequences such as Factor Xa (see, for example, Ottavi, Biochimie, 80: 289-93 (1998)), the recognition portion of suftilisin protease (see, for example, Poliak, Protein Eng. , 10: 615-19 (1997)); enterokinase (Invitrogen, San Diego, Calif.), and the like, between the translocation domain (for efficient plasma membrane expression) and the rest of the newly translated peptide can be used to facilitate purification. For example, a construct can include a polypeptide encoding a nucleic acid sequence linked to six histidine residues followed by a thioredoxin, an enterokinase cleavage site (see, for example, Williams, Biochemistry, 34: 1787-97 (1995 )), and a C-terminal translocation domain. The histidine residues facilitate detection and purification while the enteroquamsa cleavage site provides a means to purify the desired protein (s) from the remainder of the fusion protein. The technology that relates to vectors that encode fusion proteins and the application of fusion proteins are well described in the scientific and patent literature (see, for example, Kroll, DNA Cell, Biol, 12: 441-53 (1993). )). Expression vectors, either as individual expression vectors or as libraries of expression vectors, comprising the sequences encoding the ligand binding region can be introduced into a genome or into the cytoplasm or a nucleus of a cell and expressed through a variety of conventional techniques, will be described in the scientific and patent literature. See, for example, Roberts, Nature, 328: 731 (1987); Berger supra; Schneider, Protein Expr. Purif., 6435: 10 (1995); Sambrook; Tijssen; Ausubel. The product information of the biological reagent manufacturers and experimental equipment also provide information regarding known biological methods. The vectors can be isolated from natural sources, obtained from such sources as ATCC or GenBank libraries, or prepared by synthetic or recombinant methods. Nucleic acids can be expressed in expression cassettes, vectors or viruses that are stably or transiently expressed in cells (e.g., episomal expression systems). Selection markers can be incorporated into the expression cassettes and vectors to confer a selectable phenotype on the transformed cells and sequences. For example, selection markers can encode episomal maintenance and replication such that integration into the host genome is not required. For example, the label can encode antibiotic resistance (eg, chloramphenicol, kanamycin, G418, bleomycin, hygromycin) or herbicide resistance (eg, chlorosulfuron or Basta) to allow selection of those cells transformed with the desired DNA sequences (See, for example, Blondelet-Rouault, Gene, 190: 315-17 (1997), Aubrecht, J. Pharmacol. Exp. Ther., 281: 992-97 (1997)). Because selectable marker genes that confer resistance to substrates similar to neomycin or hygromycin can only be used in tissue culture, chemoresistant genes are also used as selectable markers in vitro and in vivo. A chimeric nucleic acid sequence can encode a T2R ligand binding region within any 7-transmembrane polypeptide. Because the 7-transmembrane receptor polypeptides have similar primary sequences and secondary and tertiary structures, the structural domains (eg, extracellular domain, TM domains, cytoplasmic domains, etc.) can be easily identified by sequence analysis. For example, homology modeling, Fourier analysis and helical periodicity detection can identify and characterize the seven domains with a 7-transmembrane receptor sequence. The fast Fourier transformation algorithms (FFT) can be used to estimate the dominant periods that characterize the profiles of the hydrophobicity and variability of the sequences analyzed. The increase in the detection of periodicity and the Alpha helical periodicity index could be done as by, for example, Donnelly, Protein Sci., 2: 55-70 (1993). Other alignment and modeling algorithms are well known in the art (see, for example, Peitsch, Receptors Channels, 4: 161-64 (1996); Kyte & Doolittle, J. Md. Biol., 157: 105-32 (1982); and Cronet, Protein Eng., 6: 59-64 (1993). The present invention also includes not only the nucleic acid molecules and the polypeptides having the specified chimeric and native T2R nucleic acid and amino acid sequences, but also the fragments thereof, particularly fragments of, eg, 40, 60, 80, 100, 150, 200, or 250 nucleotides, or more, as well as polypeptide fragments of, for example, 10, 20, 30, 50, 70, 100, or 150 amino acids, or more. Optionally, the nucleic acid fragments can encode an antigenic polypeptide that is capable of binding to a raised antibody against a member of the T2R family. In addition, a protein fragment of the invention can optionally be an antigenic fragment that is capable of binding to a raised antibody against a member of the T2R family. It is also contemplated that the chimeric proteins, comprising at least 10, 20, 30, 50, 70, 100, or 150 amino acids, or more, of at least one of the T2R polypeptides described herein, coupled to the additional amino acids representing all or part of another GPCR, preferably a member of the 7-transmembrane superfamily. These chimeras can be made from the present receptors and another GPCR, or they can be made by combining two or more of the present receptors. In one embodiment, a portion of the chimera corresponds to, or is derived from, the transmembrane domain of a T2R polypeptide of the invention. In another embodiment, a portion of the chimera corresponds to, or is derived from, one or more of the transmembrane regions of a T2R polypeptide described herein, and the remaining portion or portions may come from another GPCR. Chimeric receptors are well known in the art, and the techniques for creating them and the selection and boundaries of domains or fragments of G protein-coupled receptors for incorporation herein are also well known. Thus, this knowledge of those skilled in the art can easily be used to create such chimeric receptors. The use of such chimeric receptors may provide for example, a flavor selectivity characteristic of one of the receptors specifically disclosed herein, coupled with the signal transduction characteristics of another receptor, such as a well-known receptor used in the systems of the prior art. For example, a region such as a ligand binding region, an extracellular domain, a transmembrane domain, a transmembrane domain, a cytoplasmic domain, an N-terminal domain, a C-terminal domain, or any combination thereof, is can bind covalently to a heterologous protein. For example, a T2R transmembrane region can be linked to a heterologous GPCR transmembrane domain or a heterologous extracellular GPCR domain can be linked to a T2R transmembrane region. Other heterologous proteins of choice may influence, for example, green fluorescent protein, beta-galactosidase polypeptides, glutamate receptor, and rhodopsin polypeptides, for example, N-terminal fragments of rhodopsin, eg, bovine rodops. It is also within the scope of the invention to use different host cells to express the T2Rs, fragments, or variants of the invention. To obtain high levels of expression of a cloned gene or nucleic acid, such as cDNAs encoding T2Rs, fragments, or variants of the invention, one of skill typically subclones the nucleic acid sequence of interest into an expression vector containing a promoter. strong to direct transcription, a transcription / translation terminator, and if for a nucleic acid encoding a protein, a ribosome binding site for translational initiation. Suitable bacterial promoters are well known in the art and are described, for example, in Sambrook et al. Preferably, eukaryotic expression systems are used to express the subject hT2R receptor. Any of the well-known methods for introducing the above nucleotide sequences into the host cells can be used. These include the use of calcium phosphate transfection, polybrene, protoplast fusion, electroporation, liposomes, micromjection, plasma vectors, viral vectors and any of the other well-known methods for introducing cloned genomic DNA, cDNA, synthetic DNA or other foreign genetic material in a host cell (see, for example, Sambrook et al.). It is only necessary that the particular genetic engineering method used be capable of successfully introducing at least one nucleic acid molecule into the host cell capable of expressing the T2R, fragment, or variant of interest. After the expression vector is introduced into the cells, the transfected cells are cultured under conditions that favor expression of the receptor, fragment, or variant of interest, which is then recovered from the culture using standard techniques. Examples of such techniques are well known in the art. See, for example, WO 00/06593, which is incorporated by reference in a manner consistent with this description. Assays for the Detection of Compounds that Modulate the Activity of a T2R According to the Invention Methods and compositions for determining whether a test compound binds specifically to a T2R polypeptide of the invention, both in vitro and in vivo are described below. Many aspects of cell physiology can be monitored to estimate the effect of ligand binding to naturally occurring or chimeric T2Rs. These assays can be performed on intact cells expressing a T2R polypeptide, on permeabilized cells, or on membrane fractions produced by standard methods. Flavor receptors bind compounds that induce flavor and initiate the transduction of chemical stimuli into electrical signals. A G protein activated or inhibited in turn will alter the properties of target enzymes, channels, and other effector proteins. Some examples are the activation of cGMP phosphodiesterase by the transducin in the visual system, the adenylated cyclase by the stimulatory protein G, phospholipase C by Gq and other cognate G proteins, and modulation of various channels by Gi and other G proteins. The consequences of Downstream can also be examined such as the generation of diacylglycerol and IP3 by phospholipase C, and in turn, for the mobilization of calcium by IP3. The chimeric T2R polypeptides tested will typically be selected from a polypeptide having a sequence contained in SEQ ID NOS: 2 and 4 or conservatively modified fragment or variants thereof. Alternatively, the chimeric T2R proteins or polypeptides of the assay can be derived from a eukaryotic host cell, and can include a sequence of amino acids having amino acid sequence identity to SEQ ID NOs: 2 or 4 or conservatively modified variants thereof. Generally, the amino acid sequence identity will be at least 30% preferably 30-40%, more specifically 50-60, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%. Optionally, the T2R proteins or polypeptides of the assays may comprise a region of a T2R polypeptide, such as an extracellular domain, transmembrane region, cytoplasmic domain, ligand binding domain, and the like. Optionally, the T2R polypeptide, or a portion thereof, can be covalently linked to a heterologous protein to create a chimeric protein used in the assays described herein. Modulators of T2R activity can be tested using T2R proteins or polypeptides as described in the above, either recombinant or occurring naturally. The T2R proteins or polypeptides can be isolated, expressed in a cell, expressed in a membrane derived from a cell, expressed in tissue or in an animal, either recombinant or occurring naturally. For example, tongue slices, dissociated cells of a tongue, transformed cells, or membranes can be used. The modulation can be tested using one of the in vitro or in vivo assays described herein. Detection of Modulators The compositions and methods for determining whether a test compound binds specifically to a T2R receptor of the invention, both in vitro and in vivo, are described below. Many aspects of cell physiology can be monitored to estimate the effect of ligand binding to a T2R polypeptide of the invention. These assays can be performed on intact cells expressing on intact cells expressing a chemosensory receptor, on permeabilized cells or on membrane fions produced by standard or in vitro methods using de novo synthesized proteins. Flavor receptors, in vivo, bind to flavor-modulating compounds and initiate the translation of chemical stimuli into electrical signals. A G protein activated or inhibited in turn will alter the properties of target enzymes, channels, and other effector proteins. Some examples are the activation of cGMP phosphodiesterase by transducin in the visual system, adenylyl cyclase by the stimulatory protein G, phospholipase C by the Gq and other cognate G proteins, and the modulation of various channels by Gi and other G proteins. Downstream consequences can also be examined such as the generation of diacyl glycerol and IP 3 by phospholipase C, and in turn, by the mobilization of calcium by EP3. Alternatively, the T2R proteins or polypeptides of the assay can be derived from a eukaryotic host cell and can include a sequence of amino acids having amino acid sequence identity to the T2R polypeptides disclosed herein, or conservatively modified fragments or variants thereof. Generally, the amino acid sequence identity will be at least 35 to 50%, or optionally 75%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%. Optionally, the T2R proteins or polypeptides of the assays may comprise a domain of a T2R protein, such as an extllular domain, transmembrane region, transmembrane domain, cytoplasmic domains, ligand binding domain, and the like. In addition, as described above, the T2R protein or a domain thereof can be covalently linked to a heterologous protein to create a chimeric protein used in the assays described herein. Modulators of T2R receptor activity are tested using the T2R proteins or polypeptides as described above, either recombinant or occurring naturally. The T2R proteins or polypeptides can be isolated, expressed in a cell, expressed in a membrane derived from a cell, expressed in tissue or in an animal, either recombinant or occurring naturally. For example, tongue slices, dissociated cells of a tongue, transformed cells, or membranes can be used. Modulation can be tested using one of the in vitro or in vivo assays described herein. 1. In vitro Linkage Assays The taste transduction can also be examined in vitro with soluble or solid state reactions, using the T2R polypeptides of the invention. In a particular embodiment, the domains that bind ligand T2R ligand can be used in vitro in soluble reactions or in solid state for the assay to bind the ligand. It is possible that the ligand binding domain can be formed by the N-terminal domain in conjunction with additional portions of the extllular domain, such as the transmembrane domain extllular circuits. The in vitro binding assays have been used with other GPCRs, such as metabotropic glutamate receptors (see, for example, Han and Hampson, J. Biol. Chem. 274: 10008-10013 (1999)). These assays could involve the displacement of a radioactive or fluorescently labeled ligand, measuring changes in intrinsic fluorescence or changes in proteolytic susceptibility, etc. The ligand binding to a T2R polypeptide according to the invention can be tested in solution, in a bilayer membrane, optionally adhered to a solid phase, in a lipid mini-layer, or vesicles. The binding of a modulator can be tested using, for example changes in the espestrocópicas characteristics (for example, fluorescence, absorbency, refractive Index) hydrodynamic (for example, form), chromatographic or properties of solubility. In a preferred embodiment of the invention, a binding assay of [35S1GTP? S. as described above, in the activation of a GPCR, the G subunit of the G protein complex is stimulated to bind in GDP exchange for GTP. The ligand-mediated stimulation of the G protein exchange activity can be measured in a biochemical assay that measures the binding of radioactively labeled [35S1GTP? S added to protein G in the presence of a putative ligand. Typically, the membranes containing the chemosensory receptor of interest are mixed with a G protein. Potential inhibitors and / or activators and [35S1GTP? S are added to the assay, and the binding of [35s] GTP? S to the G protein is measure. The link can be measured by liquid scintillation counting or by any other means known in the art, which includes scintillation proximity assays (SPA). In other assay formats, the fluorescently labeled GTP? S can be used. 2. Fluorescence Polarization Assays In another embodiment, assays based on fluorescence polarization ("FP") can be used to detect and monitor the ligand binding. Fluorescence polarization is a versatile laboratory technique for measuring equilibrium binding, nucleic acid hybridization, and enzyme activity. Fluorescence polymerization assays are homogeneous in that they do not require a separation step such as centrifugation, filtration, chromatography, precipitation, or electrophoresis. These tests are done in real time, directly in solution and do not require any immobilized phase. The polarization values can be measured repeatedly and after the addition of reagents since the measurement of the polarization is rapid and does not destroy the sample. Generally, this technique can be used to measure the polarization values of low picomolar fluorophores at micromolar levels. This section describes how fluorescence polarization can be used in a simple and quantitative way to measure the binding of the ligands to the T2R polypeptides of the invention. When a fluorescently labeled molecule is excited by plane polarized light, it emits light that has a degree of polarization that is inversely proportional to its molecular rotation. The large fluorescently labeled molecules remain relatively stationary during the excited state (4 nanoseconds in the case of fluorescein) and the polarization of the light remains relatively constant between excitation and emission. The fluorescently small molecules rapidly break during the excited state and the polarization changes significantly between excitation and emission. Therefore, small molecules have low polarization values and large molecules have high polarization values. For example, a single-stranded fluorescein-labeled oligonucleotide has a relatively low polarization value but when hybridized to a complementary strand, it has a higher polarization value. By using FP to detect or monitor the taste-inducible compound binding that can activate or inhibit the chemosensory receptors of the invention, fluorescently labeled flavor inducing compounds or compounds that induce autofluorescence flavor can be used. The fluorescence polarization (P) is defined as: P = Mw MeSBl [Intpar + Intperp] Where Intpar is the intensity of the emission light parallel to the plane of excitation light and Intperp is the intensity of the emission light perpendicular to the plane of excitation light. P, which is a ratio of light intensities, is a number without dimension. For example, the Beacon ™ and Beacon 2000 ™. The system can be used in relation to these tests. Such systems typically express polarization in a millipolamping unit (1 Polarization Unit IMt = 1000 mP). The relationship between molecular rotation and size is described by the Perrin equation and the reader refers to Jolley, M.E. (1991) Journal of Analytical Toxicology, pp. 236-240 incorporated by reference, which gives an explanation through this equation. In summary, the Perrin equation states that the polarization is directly proportional to the rotational relaxation time, the time it takes for a molecule to rotate through an angle of approximately 68.5 °. The rotational relaxation time is relative to the viscosity (eta.), Absolute temperature (T), molecular volume (V), and gas constant (R) by the following equation: 2 (Rotational Relaxation Time) = 3 V RT . The rotational relaxation time is small (~ nanosecond) for small molecules (eg fluorescein) and large (~ 100 nanoseconds) for large molecules (eg immunoglobulins). If the viscosity and temperature are kept constant, the rotational relaxation time, and therefore the polarization, is directly related to the molecular volume. Changes in molecular volume may be due to interactions with other molecules, disassociation, polymerization, degradation, hybridization, or adaptive changes of the fluorescently labeled molecule. For example, fluorescence polarization has been used to measure the enzymatic cleavage of large fluorescein-labeled polymers by proteases, DNases, and RNases. It has also been used to measure the equilibrium bond for protein / protein interactions, antibody / antigen binding, and protein / DNA binding. A. Solid and Soluble Solid State High Performance Assays In still another embodiment, the invention provides soluble assays utilizing a T2R polypeptide; or a cell or tissue that expresses a T2R polypeptide. In another embodiment, the invention provides in vitro assays based on solid phase in a high throughput format, wherein the T2R polypeptide, or cell or tissue expressing T2R polypeptide, adhere to a solid phase substrate or a flavor-stimulating compound. and is contacted with a T2R receptor, and on a detected link using an appropriate tag or antibody raised against the T2R receptor. In the high throughput assays of the invention, it is possible to sort several thousand different modulators or ligands in a single day. In particular, each cavity of a microtiter plate can be used to run a separate assay against a selected potential modulator, or, if the effects of concentration or incubation time are to be observed, each 5-10 cavities can test a single modulator . Thus, a single standard microtiter plate can analyze approximately 100 modulators (eg, 96). If the plates of 1536 cavities are used, then a single plate can easily analyze from about 1000 to about 1500 different compounds. It is also possible to analyze multiple compounds in each well of the plate. It is possible to analyze several different plates per day; Test classifications for up to about 6,000-20,000 different compounds is possible using the integrated systems of the invention. More recently, microfluidic procedures for the handling of the reagent have been developed. The molecule of interest can be linked to the component in the solid state, directly or indirectly, via the covalent or non-covalent bond, for example, by way of a brand. The brand can be any of a variety of components. In general, a molecule that binds the label (a branded binder) is attached to a solid support, and the labeled molecule of interest (e.g., the flavor transducing molecule of interest) is attached to the solid support by the interaction of the brand and the brand binder. A number of brands and brand binder can be used, based on known molecular interactions well described in the literature. For example, when a brand has a natural binder, for example, biotin, protein A, or protein G, they can be used in conjunction with appropriate brand binders (avidin, streptavidin, neutravidin, the Fc region of an immunoglobulin, etc.). . Antibodies to molecules with natural binders such as biotin are also widely available and appropriate brand binders (see, SIGMA Immunochemicals 1998 catalog SIGMA, St. Louis Mo.). Similarly, any gene or antigenic compound can be used in combination with an appropriate antibody to form a brand / brand binder pair. Thousands of specific antibodies are commercially available and many additional antibodies are described in the literature. For example, in a common configuration, the label is a first antibody and the label binder is a second antibody that recognizes the first antibody. In addition to antibody-antigen interactions, receptor-ligand interactions are also appropriate as brands and brand binder pairs. For example, agonists and antagonists of cell membrane receptors (e.g., receptor-cell ligand interactions such as transferin, c-kit, viral receptor ligands, cytosine receptors, chemokine receptors, methlecuquin receptors, receptors of immunoglobulm, and antibodies, the caderein family, the mtegrin family, the selectin family and the like; see, for example, Pigott & Power, The Adhesion Molecule Facts Book I (1993)). Similarly, toxins and poisons, viral epitopes, hormones (for example, opiates, spheroids, etc.), intracellular receptors (for example, that mediate the effect of several small ligands, which include spheroids, thyroid hormone, retmoids and vitamin D; peptides), drugs, lectins, sugars, nucleic acids (both linear and cyclic polymer configurations), oligosaccharides, proteins, phospholipids and antibodies can all interact with several cellular receptors. Synthetic polymers, such as polyurethanes, polyesters, polycarbonates, polyureas, polyamides, polyethylene imines, polyaplene sulfoxides, polysiloxanes, polyamides, and polyacetates can also form a suitable brand or binder. Many other branded brand / binder pairs are also useful in assay systems described herein, as would be apparent to one of skill in reviewing this disclosure. Common linkers such as peptides, polyethers, and the like can also serve as labels, and include approximately 5 and 200 amino acid polypeptide sequences. Such flexible linkers are known to persons of skill in the art. For example, poly (ethylene glycol) linkers are available from Shearwater Polimers, Inc. Huntsville, Ala. These linkers optionally have amide bonds, sulfhydryl bonds or heterofunctional linkages. Brand links are fixed to solid substrates using any of a variety of currently available methods. Solid substrates are commonly derived or functionalized by exposing all or a portion of the substrate to a chemical reagent that attaches a chemical group to the surface that is reactive with a branching linker portion. For example, groups that are suitable to adhere to a longer chain portion would include amines, hydroxyl, thiol, and carboxyl groups. The ammoalkylsilanes and hydroxyalkyl silanes can be used to functionalize a variety of surfaces, such as glass surfaces. The construction of such solid phase biopolymer arrays are well described in the literature. See, for example, Merrifield, J. Am. Chem. Soc, 85: 2149-2154 (1963) (describing solid phase synthesis of, for example, peptides); Geysen et al., J. Immun. Meth., 102: 259-274 (1987) (describing synthesis of solid phase components or tips); Frank & Dopng, Tetrahedron, 44: 60316040 (1988) (describing synthesis of various peptide sequences on cellulose discs); Fodor et al., Science, 251: 767-777 (1991); Sheldon et al., Climcal Chemistry, 39 (4): 718-719 (1993); and Kozal et al., Nature Medicine, 2 (7): 753-759 (1996) (describing all arrays of biopolymers fixed to solid substrates). Non-chemical methods for fixing the brand bonds to substrates include other common methods, such as heat, UV crosslinking and the like. 3. Cell-based Assays In a preferred embodiment, a T2R protein is expressed in a eukaryotic cell either in unmodified forms or as variant or truncated, chimeric receptors with or preferably without an accompanying, heterologous sequence that facilitates its maturation and that is directs through the secretory route. Such T2R polypeptides can be expressed in any eukaryotic cell, such as HEK-293 cells. Preferably, the cells comprise a functional G protein, for example, Ga? 5, or a G. a? 6, chimeric, gustucin or transducin or a chimeric G protein such as G16gust44 which is capable of coupling the chimeric receptor to a protein pathway. metacellular signaling or to a signaling protein such as phospholipase C. Activation of T2R receptors in such cells can be detected using any standard method, such as by detecting changes in intracellular calcium by detecting FURA-2 dependent fluorescence in the cell. Such an essay is the basis of the experimental discoveries presented in this application. Frequently activated GPCR receptors are substrates for kinases that phosphores the C-terminal tail of the receptor (and possibly other sites as well). A) Yes, activators will promote the 32P transfer of radiolabeled ATP to the receptor, which can be analyzed with a scintillation counter. The phosphorylation of the C-terminal tail will promote the binding of the arrestin-like proteins and will interfere with the binding of the G proteins. For a general review of the transduction signal will be GPCR and methods to analyze the signal transduction see, for example, Methods in Enzymology, vols. 237 and 238 (1994) and volume 96 (1983); Bourne et al., Nature, 10: 349: 117-27 (1991); Bourne et al., Nature, 348: 125-32 (1990); Pitcher and collaborators, Annu. Rev. Biochem., 67: 653-92 (1998). The T2R modulation can be analyzed by comparing the response of the chimeric T2R polypeptides according to the invention treated with a putative T2R modulator to the response of an unreacted control sample or a sample containing a known "positive" control. Such putative T2R modulators can include molecules that either inhibit or activate the activity of the T2R polypeptide. In one embodiment, control samples treated with a compound that activates T2R are assigned a relative T2R activity value of 100. Inhibition of a T2R polypeptide is achieved when the value of the T2R activity relative to the control sample is of about 90%, optionally 50%, optionally 25-0%. Activation of a T2R polypeptide is achieved when the T2R activity value relative to the control is 110%, optionally 150%, 200-500%, or 1000-2000%. Changes in ion flux can be estimated by determining changes in ion polymerization (ie, electrical potential) of the cell or membrane that expresses a T2R polypeptide. A means to determine changes in cell polarization is by measuring changes in current (thus measuring changes in polarization) with voltage fixation and patch fixation techniques (see, for example, the "attached" mode). the cell, the "inside-out" mode, and the "whole cell" mode, for example, Ackerman et al., New Engl. J Med., 336: 1575-1595 (1997)). Whole cell currents are conveniently determined using the standard. Other known assays include: radiolabelled ion flow assays and fluorescence assays using voltage sensitive dyes (see, for example, Vestergarrd-Bogind et al., J. Membrane Biol., 88: 67-75 (1988); Gonzales &; Tsien, Chem. Biol., 4: 269-277 (1997), Daniel et al., J. Pharmacol. Meth., 25: 185-193 (1991); Holevinsky et al., J. Membrane Biology, 137: 59- 70 (1994)).

The effects of the test compounds on the function of the polypeptides can be measured by examining any of the parameters described in the above. Any suitable physiological change affecting the GPCR acty can be used to estimate the influence of a test compound on the polypeptides of this invention. When the functional consequences are determined using intact cells or animals, a variety of effects can also be measured such as transmitter release, hormone release, transcpptional changes to both known and characterized genetic markers (eg, northern spotting), changes in cellular metabolisms, such as cell growth or pH changes and changes in intracellular second messengers such as Ca.2 +, IP3, cGMP, or cAMP. Preferred assays for GPCRs include cells that are loaded with ion-sensitive dyes or voltage to report receptor acty. Assays for determining the acty of such receptors can also use known agonists and antagonists for other receptors coupled to G proteins as controls to estimate the acty of the tested compounds. In assays to identify modulating compounds (eg, agonist, antagonists), changes in the ion level the cytoplasm or membrane voltage will be monitored using a fluorescent indicator of ion- or membrane-sensitive voltage, respectively. Among the indicators sensitive to ions and voltage probes that can be used are those that are published in the Molecular Probes 1997 Catalog. For receptors coupled to protein G, promiscuous G proteins such as Gai5 and Gai6 can be used in the lesson assay (Wilkie et al., Proc. Natd Acad. Sci., 88: 10049-10053 (1991)). Alternatively, other G proteins such as gustducma, transducin and chimeric G proteins such as Gal6gust44 or G16g44 can be used. The receptor activation initiates subsequent intracellular events, for example, increases in the second messengers. Activation of some G protein-coupled receptors stimulate the formation of nositol triphosphate (IP3) through phospholipase C-mediated hydrolysis of phosphatidylinositol (Berpdge &Irvine, Nature, 312: 315-21 (1984)). IP3 in turn stimulates the release of intracellular calcium ion stores. Thus, a change in the cytoplasmic calcium ion levels, or a change in the levels of the second messenger such as IP3 can be used to estimate the receptor function coupled to protein G cells expressing such G protein-coupled receptors can exhibit levels of cytoplasmic calcium increased as a result of the contribution of both calcium release from intracellular stores and extracellular calcium entering via the plasma membrane ion channels. In a preferred embodiment, the T2R polypeptide acty is measured by expressing the T2R gene in a heterologous cell with a promiscuous G protein that binds the receptor to a phospholipase C signal transduction pathway (see Offermanns &Simon, J. Biol. Chem., 270: 15175-15180 (1995)). Preferably, the cell line is HEK-293 (which does not normally express T2R genes) and the promiscuous G protein is Gai5 (Offermanns &Simon, supra) or a chimeric G protein such as Gal6gust44. The modulation of the test transduction is analyzed by measuring changes in the cell-cell Ca2 + levels, which change in response the modulation of the transduction path of the T2R signal via the administration of a molecule that is associated with the polypeptide. T2R. Changes in Ca2 + levels are optionally measured using fluorescent Ca2 + indicator dyes and fluorometric imaging. In another embodiment, the hydrolysis of phosphatidyl inositol (Pl) can be analyzed according to US Pat. No. 5, 436.128, incorporated herein by reference. Briefly, the assay involves labeling the cells with 3H-m? O? Us? Tol for 48 hours or more. The labeled cells are treated with a test compound for 1 hour. The treated cells are lysed and extracted into chloroform-methanol-water after the inositol phosphates were prepared by ion exchange chromatography was quantified by scintillation counting. Fold stimulation is determined by calculating the ratio of CPM in the presence of agonist, to CPM control buffer. Similarly, folding inhibition is determined by calculating the ratio of CPM in the presence of the antagonist, to CPM in the presence of buffer control (which may or may not contain an agonist). Other receptor assays that may involve determining the level of the intracellular cyclic nucleotides, for example, cAMP or cGMP. In cases where the activation of the receptor results in a decrease in the levels of the cyclic nucleotides, it may be preferable to expose the cells to people who increase the levels of the cyclic nucleotide nucleotides, for example, forskolin, prior to the addition of a compound of activation of the receptor to the cell in the assay. In one embodiment, changes in intracellular cAMP or cGMP can be measured using immunoassays. The method described in Offermanns &; Simon, J. Bio. Chem., 270: 15175-15180 (1995), can be used to determine the level of cAMP. Also, the method described in Felley-Bosco et al., Am. J. Resp. Cell and Mol. Biol., 11: 159-164 (1994), can be used to determine the level of cGMP. In addition, a test kit for measuring cAMP and / or cGMP is described in U.S. Patent No. 4,115,538, incorporated herein by reference. In another embodiment, transcript levels can be measured to estimate the effects of a test compound on signal transduction. A host cell containing the T2R polypeptide of interest is contacted with a test compound for a sufficient time to effect any of the interactions and then the level of gene expression is measured. The amount of time to perform such interactions can be determined empirically, such as by running a time course and measuring the level of transcription as a function of time. The amount of transcription can be measured by using any method known to those of skill in the art to be suitable. For example, mRNA expression of the protein of interest can be detected using northern blots or its polypeptide products can be identified using immunoassays. Alternatively, the assays based on No. 5,436,128, incorporated herein by reference. Reporter genes can be, for example, chloramphenicol acetyltransferase, luciferase, beta-galactosidase, beta-lactamase and alkaline phosphatase. In addition, the protein of interest can be used as an indirect reporter via the binding to a second reporter such as the green fluorescent protein (see, for example, Mistili &Spector, Nature Biotechnology, 15: 961-964 (1997 )). The amount of the transcript is then compared to the amount of transcription in either the same cell in the absence of the test compound, or it can be compared to the amount of transcription in a substantially identical cell lacking the T2R polypeptide (s) of interest. A substantially identical cell can be derived from the same cells from which the recombinant cell was prepared but which would not have been modified by the introduction of heterologous DNA. Any difference in the amount of the transcript indicates that the test compound has somehow altered the activity of the T2R polypeptide of interest. 4. Transgenic Non-Human Animals Expressing Chemosensory Receptors Non-human animals expressing one or more flavor receptor sequences of the invention can also be used for receptor assays. Such an expression can be used to determine whether a test compound binds specifically to a mammalian taste transmembrane receptor complex in vivo by contacting a stable or transiently transfected non-human animal with nucleic acids encoding chemosensory receptors or protein binding regions. ligand thereof with a test compound and determine if the animal reacts to the test compound by binding specifically to the receptor polypeptide complex. Animals transfected or infected with the vectors of the invention are useful particularly for assays to identify and characterize flavor stimuli that can bind to a specific or set of receptors. Such vector-infected animals expressing human taste receptor sequences can be used for in vivo classification of flavor stimuli and their effect on, for example, cell physiology (eg, on flavor neurons), on the CNS, or behavior. Means for infecting / expressing nucleic acids and vectors, either individually or as libraries, are well known in the art. A variety of individual cells, organ or whole animal parameters can be measured by a variety of means. The T2R sequences of the invention can be expressed, for example, in animal test tissues by delivery with an infecting agent, for example, adenovirus expression vector. The endogenous taste receptor genes may remain functional and wild-type (native) activity may still be present. In other situations, where it is desirable that all taste receptor activity be by the introduced exogenous hybrid receptor, the use of a blocking line is preferred. Methods for the construction of non-human transgenic animals, particularly transgenic mice, and the selection and preparation of recombinant constructs to generate transformed cells are well known in the art. The construction of a "blocked" and animal cell is based on the premise that the level of expression of a particular gene in a mammalian cell can be diminished or completely invalidated by introducing a new DNA sequence into the genome that interrupts some portion of the DNA sequence of the gene to be deleted. Also, "gene trap insertion" can be used to interrupt a host gene and mouse embryonic stem cells (s) can be used to produce transgenic blocking animals (see, eg, Holzschu, Transgenic Res 6: 97- 106 (1997)). The exogenous insertion is typically by homologous recombination between complementary nucleic acid sequences. The exogenous sequence is some portion of the target gel to be modified, such as exonic, intronic or transcriptional regulatory sequences, or any genomic sequence that is capable of affecting the level of expression of the target gene: or a combination thereof. The gene that is directed through the pathway of homologous recombination in pluripotent embryonic stem cells allows to precisely modify the genomic sequence of interest. Any technique can be used to create, classify, propagate, a blocking animal, for example, see Bi voet, Hum. Mol. Genet 7: 53-62 (1998); Moreadith, J. Mol. Med. 75: 208-216 (1997); Tojo, Cytotechnology 19: 161-165 (1995); Mudgett, Methods Mol. Biol. 48: 167-184 (1995); Longo, Transgenic Res. 6: 321-328 (1997); U.S. Patent Nos. 5,616,491; 5,464,764; 5,631,153; 5,487,992; 5,627,059; 5,272,071; WO 91/09955; WO 93/09222; WO 96/29411; WO 95/31560; WO 91/12650. The nucleic acids of the invention can also be used as reagents to produce "blocking" human cells and their progeny. Likewise, the nucleic acids of the invention can also be used as reagents to produce "deactivations" in mice. The sequences of the human or rat T2R gene can replace the orthologous T2R in the mouse genome. In this way, a mouse expressing a human or rat T2R is produced. This mouse can then be used to analyze the function of the human or rat T2Rs, and to identify ligands for such T2Rs. Modulators The compounds tested as modulators of a member of the T2R family can be any small chemical compound, or a biological entity, such as a protein, sugar, nucleic acid or lipid. Alternatively, the modulators may be genetically altered versions of a member of the T2R family. Typically, the test compounds can be small chemical molecules and peptides. Essentially any chemical compound can be used as a potential modeler or ligand in the assays of the invention, although most frequently compounds can be dissolved in aqueous or organic solutions (especially based on DMS) are used. The assays can be designed to classify large chemical libraries by automating the testing steps and providing compounds from any convenient source to the assays, which are typically run in parallel. For example, in microtiter formats or microtiter plates in robotic tests). It will be appreciated that there are many suppliers of chemical compounds, including Sigma (St. Louis, Mo.), Aldrich (St. Louis, Mo.), Sigma-Aldrich (St. Louis, Mo.), Fluka Chemika-Biochemica Analytika (Buchs). , Switzerland) and the like. In one embodiment, high throughput classification methods involve providing a combination chemical or peptide library containing a large number of potential therapeutic compounds (potential modulator or ligand compounds). Such "combination chemical libraries or" ligand libraries "are then classified in one or more assays, as described herein, to identify those library members (particular chemical species or subclasses) that exhibit a desired characteristic activity. thus identified can serve as conventional "guide compounds" or can themselves be used as current potential or consumer products A combination chemical library is a collection of diverse chemical compounds generated by either chemical synthesis or biological synthesis, by combining a number of chemical "building blocks", such as reagents For example, a linear combination chemical library such as a polypeptide library is formed by combining a set of chemical building blocks (amino acids) in a possible manner for a length of given compound (that is, the number of amino acids in a given polypeptide post.) Millions of chemical compounds can be synthesized through such a combination mixture of chemical building blocks. The preparation and classification of chemical combination libraries are well known to those of skill in the art. Such combination chemical libraries include, but are not limited to, peptide libraries (see, for example, U.S. Patent No. 5,010,175, Furka, Int. J. Pept. Prot. Res., 37: 487-93 (1991 ) and Houghton et al., Nature, 354: 84-88 (1991)). Other chemistries to generate chemical diversity libraries can also be used. Such chemistries include, but are not limited to: peptoids (e.g., WO 91/19735), encoded peptides (e.g., WO 93/20242), random bio-oligomers (e.g., WO 92/00091), benzodiazepines (e.g. US Pat. No. 5,288,514), funders such as hydantoins, benzodiazepines and dipeptides (Hobbs et al., PNAS., 90: 6909-13 (1993)), polypeptides (Hagihara et al., J. Amer. Chem. Soc, 114: 6568 (1992)), non-peptide peptidomimetics with glucose structure (Hirschmann et al., J. Amer. Chem. Soc, 114: 9217-18 (1992)), analogous organic synthesis of small compound libraries (Chen et al., J. Amer. Chem. Soc, 116: 2661 (1994)), oligocarbamates (Cho et al., Science, 261: 1303 (1993)), peptidyl phosphonates (Campbell et al., J. Org. Chem., 59: 658 (1994)). ), nucleic acid libraries (Ausubel, Berger, and Sambrook, all supra), libraries of peptide nucleic acid (US Pat. No. 5,539,083), bookstores of antibodies (Vaughn et al., Nature Biotechnology, 14 (3): 309-14 (1996) and PCT / US96 / 10287), carbohydrate libraries (Liang et al., Science, 274: 1520-22 (1996) and the patent No. 5,593,853), libraries of small organic molecules (benzodiazepines, Baum, C & amp; amp;; EN, January 18, page 33 (1993); thiazolidinones and metathiazanones, U.S. Patent No. 5,549,974; pinrolidines, U.S. Patent Nos. 5,525,735 and 5,519,134; morpholmo compounds, U.S. Patent No. 5,506,337; benzodiazepines, 5,288,514, and the like). Devices for the preparation of combination libraries are commercially available, see, for example, 357 MPS, 390 MPS (Advanced Chem Tech, Louisville Ky.), Symphony (Rainm, Woburn, Mass.), 433A (Applied Biosystems, Foster City, Calif.), 9050 Plus (Millipore, Bedford, Mass.)). In addition, numerous combination libraries are themselves commercially available (see, for example, ComGenex, Princeton, NJ; Tppos, Inc., St. Louis, Mo .; 3D Pharmaceuticals, Exton, Pa .; Martek Biosciences; Columbia, Md., Etc.). In one aspect of the invention, the T2R modulators can be used in any food product, confectionery, pharmaceutical composition, or ingredient thereof by thereby modulating the taste of the product, composition, or ingredient in a desired manner. For example, T2R modulators that induce the sweet taste sensation or umami can be added to provide an improved sweet or umami flavor to a product or composition while the T2R modulators that increase sweet taste sensations or umami can be added to increase the sweet or umami flavor of another compound in a composition such as a food or beverage food product or composition. Also, the invention provides means to identify sweet or umami compounds and enhancers found in foods, beverages and medicinal substances and that produce improved flavor foods, beverages and medicinal substances lacking or having a reduced amount thereof. Use of Compounds Identified by the Invention Compounds identified according to the invention can be added to foods, beverages or medicinal compositions to modulate, preferably block the bitter taste triggered by the activation of hT2R8, hTlR14, and hT2R54 by the bitter compounds, for example , chlorogenic lactones found in coffee and other foods and beverages and structurally related and, for example, other bitter chlorogenic compounds. For example, compounds that block the activation of hT2R8, hT2R14 or hT2R54 by chlorogenic lactones or related compounds can be used as addictives in coffee beverages and coffee flavored beverages in order to block the bitter taste associated with chlorogenic lactones. or other bitter compounds. For example, these compounds can be added to coffee flavor foods and beverages in an amount effective to inhibit bitter taste. As previously noted, preferably, the flavor modulating properties of the compounds identified in the cell-based assays will be confirmed in taste tests, eg, human taste tests. Equipment The subject chimeric T2R genes and their homologs are useful tools for identifying taste receptor cells, for forensic and paternity determinations, and for examining flavor transduction. Reagents specific to the T2R family member that specifically hybridize to T2R nucleic acids such as T2R probes and primers and T2R specific reagents that specifically bind to a T2R protein, eg, T2R antibodies, are used to examine cell expression flavor cell and taste transduction regulation. Nucleic acid assays for the presence of DNA and RNA for a member of the T2R family in a sample include numerous techniques that are known to those skilled in the art, such as southern analysis, northern analysis, spot spotting, protection of RNase, SI analysis, amplification techniques such as PCR, and in situ hybridization. In in situ hybridization, for example, the target nucleic acid is released from its cellular surroundings so as to be available for hybridization within the cell while preserving the cell morphology for subsequent interpretation and analysis. The following articles provide an overview of the technique of in situ hybridization: Singer et al., Biotechniques, 4: 230250 (1986); Haase et al., Methods in Virology, vol. VII, 189-226 (1984); and Ñames et al., eds., Nucleic Acid Hybridization: A Practical Approach (1987). In addition, a T2R protein can be detected with the various immunoassay techniques described in the foregoing. The test sample is typically compared to either a positive control (e.g., a sample expressing a recombinant T2R protein) or a negative control. The present invention also provides equipment for the classification of modulators of members of the T2R family. Such equipment can be prepared from readily available materials and reagents. For example, such kits may comprise any one or more of the following materials: T2R nucleic acids or proteins, reaction tubes, and instructions for testing T2R activity. Optionally, the kit contains a functional T2R polypeptide. A wide variety of equipment and components can be prepared according to the present invention, depending on the proposed user of the equipment and the particular needs of the user. Having now generally described the invention, it will be more readily understood by reference to the following examples, which are provided by way of illustration and are not intended to be limiting. It is understood that various modifications and changes can be made to the exemplary embodiments disclosed herein without departing from the spirit and scope of the invention. EXAMPLES Example 1 In this example, the inventors show that 3CoQAL, a bitter chlorogenic lactone compound, specifically activates human bitter receptors hT2R8, hT2R14, and hT2R54 having the DNA sequence contained in this application. Activation of these receptors by 3CoQAL is measured in a cell-based assay that detects changes in intracellular calcium concentration. Briefly, human embryonic kidney cells are seeded in 48 cavity tissue culture plates. 24 hours later, the cells are transiently transfected with the plasmid containing either the hT2R8 nucleic acid sequence, hT2R14 or hT2R54, and the plasmid containing a chimeric G protein (G16gust44). Another 24 hours later, the cells are incubated with a fluorescent dye specific for calcium (Fluo-4 or Fura-2; Molecular Probes) that provides a simple and reliable fluorescence-based method to detect changes in calcium concentration within the cell. Activation of the T2Rs induces a signaling shell that drives PLC activation and a subsequent increase in intracellular calcium concentration. This increases in calcium concentration changes to the fluorescence properties of the calcium dye within the cells. These changes are monitored using fluorescence microscopy and specific design software (Imaging Workbench, Axon). Using this procedure it was observed that both the 3CQAL and 4CQAL chlorogenic lactones specifically activate the cells (increase the intracellular calcium concentration) that these T2Rs express (See Figure 3). (See proven list of dosages in Figure 2). Example 2 In this example, the inventors show that 3CQAL and 4CQAL, two bitter chlorogenic lactone compounds, specifically activate the human bitter receptors, hT2R8, hT2R14 and hT2R54, which have the DNA sequence contained in SEQ ID NO: 2, 4 and 6 respectively in this application. Activation of this receptor by 3CoQAL and 4CoQAL is measured in a cell-based assay that detects changes in intracellular calcium concentration. Briefly, human embryonic kidney cells are seeded in 48 cavity tissue culture plates. 24 hours later, the cells are transiently transfected with the plasmid containing either the nucleic acid sequence hT2R8, hT2R14 or hT2R54, and the plasmid containing a chimeric G protein (G16gust44). Another 24 hours later, the cells are incubated with a fluorescent dye specific for calcium (Fluo-4 or Fura-2).; Molecular Probes) that provides a simple and reliable, rapid fluorescence based method to detect changes in calcium concentration within the cell. Activation of the T2Rs induces a signaling cascade that leads to PLC activation and a subsequent increase in intracellular calcium concentration. This increases in the calcium concentration changes to the fluorescence properties of the calcium dye within the cells. These changes are monitored using fluorescence microscopy and specific design software (Imaging Workbench, Axon). Using this procedure it was observed that both of the chlorogenic lactones 3CQAL and 4CQAL specifically activate the cells (increases the intracellular calcium concentration) expressed by these T2Rs (See Figure 3). Example 3 In this example, the inventors show that 4FQAL, a bitter chlorogenic lactone compound, specifically activates human bitter receptors, hT2R8, hT2R14 and hT2R54, having the DNA sequence contained in SEQ ID NO: 2, 4 and 6 respectively in this application. Activation of this receptor by 4QFQAL is measured in a cell-based assay that detects changes in intracellular calcium concentration. Briefly, human embryonic kidney cells are seeded in 48 cavity tissue culture plates. 24 hours later, the cells are transiently transfected with the plasmid containing either the hT2R8 nucleic acid sequence, hT2R14 or hT2R54, and the plasmid containing a chimeric G protein (G16gust44). Another 24 hours later, the cells are incubated with a fluorescent dye specific for calcium (Fluo-4 or Fura-2; Molecular Probes) that provides a rapid, simple and reliable fluorescence-based method to detect changes in the calcium concentration inside the cell. Activation of the T2Rs induces a signaling cascade that leads to PLC activation and a subsequent increase in intracellular calcium concentration. This increases in the calcium concentration changes to the fluorescence properties of the calcium dye within the cells. These changes are monitored using fluorescence microscopy and specific design software (Imaging Workbench, Axon). Using this procedure the inventors observed that both of the 4FQAL chlorogenic lactones specifically activate the cells (increase the intracellular calcium concentration) expressed by these T2Rs.

(See Figure 4). SEQUENCES OF HT2R GENES AND POLYPEPTIDES EXAMPLES IN THE I PRESENTED Nucleotide > hT2R8 (SEQ ID NO: 2) ATGTTCAGTCCTGCAGATAACATCTTTATAATCCTAATAACTGGAGAATTCATACTAGGAATATTGGGGAATGGA TACATTGCACTAGTCAACTGGATTGACTGGATTAAGAAGAAAAAGATTTCCACAGTTGACTACATCCTTACCAAT TTAGTTATCGCCAGAATTTGTTTGATCAGTGTAATGGTTGTAAATGGCATTGtAATAGTACTGAACCCAGATGTT TATACAAAAAATAAACAACAGATAGTCATTTTTACCTTCTGGACATTTGCCAACTACTTAAATATGTGGATTACC ACCTGCCTTAATGTCTTCTATTTTCTGAAGATAGCCAGTTCCTCTCATCCACTTTTTCTCTGGCTGAAGTGGAAA ATTGATATGGTGGTGCACTGGATCCTGCTGGGATGCTTTGCCATTTCCTTGTTGGTCAGCCTTATAGCAGCAATA GTACTGAGTTGTGATTATAGGTTTCATGCAATTGCCAAACATAAAAGAAACATTACTGAAATGTTCCATGTGAGT] 0 AAAATACCATACTTTGAACCCTTGACTCTCTTTAACCTGTTTGCAATTGTCCCATTTATTG GTCACTGATATCA TTTTTCCTTTTAGTAAGATCTTTATGGAGACATACCAAGCAAATAAAACTCTATGCTACCGGCAGTAGAGACCCC AGCACAGAAGTTCATGTGAGAGCCATTAAAACTATGACTTCATTTATCTTCTTTTTTTTCC ATACTATATTTCT TCTATTTTGATGACCTTTAGCTATCTTATGACAAAATACAAGTTAGCTGTGGAGTTTGGAGAGATTGCAGCAATT CTCTACCCCTTGGGTCACTCACTTATTTTAATTGTTTTAAATAATAAACTGAGGCAGACATTTGTCAGAATGCTG ACATGTAGAAAAATTGCCTGCATGATATGA Protein > hT2R8 (SEQ ID NO: 3) MFSPADNIFIILITGEFILGILGNGYIALVN IDWIKKKKISTVDYILTN VIARICLISV WNGIVIV NPDVYTKNKQQIVIFTF TFANYL M IT TCL VFYFLKIASSSHP FLW K KIDMWH I GCFAISL VS IAAI VLSCDYRFHAIAKHKRNITEMFHVSKIPYFEP TLFNLFAIVPFIVSLIS FFLLVRSL RHTKQIKLYATGSRDPSTEVHVRAIKTMTSFIFFFFLYYIS SILMTFSY MTKYK AVEFGEIAAILYPLGHSLI IV NNKLRQTFVRML TCRK1ACMI Nucleotide > hT2R14 (SEQ ID NO: 4) -n ATGGGTGGTGTCATAAAGAGCATATTTACATTCGTTTTAATTGTGGAATTTATAATTGGAAATTTAGGAAATAGT TTCATAGCACTGGTGAACTGTATTGACTGGGTCAAGGGAAGAAAGATCTCTTCGGTTGATCGGATCCTCACTGCT TTGGCAATCTCTCGAATTAGCCTGGTTTGGTTAATATTCGGAAGCTGGTGTGTGTCTGTGTTTTTCCCAGCTTTA TTTGCCACTGAAAAAATGTTCAGAATGCTTACTAATATCTGGACAGTGATCAATCATTTTAGTGTCTGGTTAGCT ACAGGCCTCGGTACTTTTTATTTTCTCAAGATAGCCAATTTTTCTAACTCTATTTTTCTCTACCTAAAGTGGAGG GTTAAAAAGGTGGTTTTGGTGCTGCTTCTTGTGACTTCGGTCTTCTTGTTTTTAAATATTGCACTGATAAACATC CATATAAATGCCAGTATCAATGGATACAGAAGAAACAAGACTTGCAGTTCTGATTCAAGTAACTTTACACGATTT TCCAGTCTTATTGTATTAACCAGCACTGTGTTCATTTTCATACCCTTTACTTTGTCCCTGGCAATGTTTCTTCTC CTCATCTTCTCCATGTGGAAACATCGCAAGAAGATGCAGCACACTGTCAAAATATCCGGAGACGCCAGCACCAAA GCCCACAGAGGAGTTAAAAGTGTGATCACTTTCTTCCTACTCTATGCCATtTTCTCTCTGTCTTTTTTCATATCA GTTTGGACCTCTGAAAGGTTGGAGGAAAATCTAATTATTCTTTCCCAGGTGATGGGAATGGCTTATCCTTCATGT CACTCATGTGTTCTGATTCTTGGAAACAAGAAGCTGAGACAGGCCTCTCTGTCAGTGCTACTGTGGCTGAGGTAC ATGTXCAAAGATGGGGAGCCCTCAGGTCACAAAGAATTTAGAGAATCATCTTGA Protein > hT2R14 (SEQ I D NO: 5) MGGVIKSIFTFV IVEFIIGNLGNSFIALVNCIDWVKGRKISSVDRI TA LAISRISLVWLIFGSWCVSVFFPALFATEKMFRM TNI TVI HFSV A TGLGTFYF KIANFSNSIFLYLKMRVKKVV VLLLVTSVF FLNIAl.INI HINASINGYRRNKTCSSDSSNFTRFSSLIV TSTVFIFIPFT SLAMFL IFSMWKHRKKMQHTVKISGDASTKAHRGVKSVITFFL YAIFSLSFFIS VWTSER EENLIILSQVMGMAYPSCHSCVLI GNKKLRQASLSVLLW RY FKDGEPSGHKEFRESS Nucleotide > hT2R54 (SEQ ID NO: 6) ATGACTAAACTCTGCGATCCTGCAGAAAGTGAATTGTCGCCATTTCTCATCACCTTAATTTTAGCAGTTTTACTT GCTGAATACCTCATTGGTATCATTGCAAATGGTTTCATCATGGCTATACATGCAGCTGAATGGGTTCAAAATAAG GCAGTTTCCACAAGTGGCAGGATCCTGGTTTTCCTGAGTGTATCCAGAATAGCTCTCCAAAGCCTCATGATGTTA GAAATTACCATCAGCTCAACCTCCCTAAGTTTTTATTCTGAAGACGCTGTATATTATGCATTCAAAATAAGTTTT ATATTCTTAAATTTTTGTAGCCTGTGGTTTGCTGCCTGGCTCAGTTTCTTCTACTTTGTGAAGATTGCCAATTTC TCCTACCCCCTTTTCCTCAAACTGAGGTGGAGAATTACTGGATTGATACCCTGGCTTCTGTGGCTGTCCGTGTTT ATTTCCTTCAGTCACAGCATGTTCTGCATCAACATCTGCACTGTGTATTGTAACAATTCTTTCCCTATCCACTCC TCCAACTCCACTAAGAAAACATACTTGTCTGAGATCAATGTGGTCGGTCTGGCTTTTTTCTTTAACCTGGGGATT GTGACTCCTCTGATCATGTTCATCCTGACAGCCACCCTGCTGATCCTCTCTCTCAAGAGACACACCCTACACATG GGAAGCAATGCCACAGGGTCCAACGACCCCAGCATGGAGGCTCACATGGGGGCCATCAAAGCTATCAGCTACTTT CTCATTCTCTACATTTTCAATGCAGTTGCTCTGTTTATCTACCTGTCCAACATGTTTGACATCAACAGTCTGTGG AATAATTTGTGCCAGATCATCATGGCTGCCTACCCTGCCAGCCACTCAATTCTACTGATTCAAGATAACCCTGGG CTGAGAAGAGCCTGGAAGCGGCTTCAGCTTCGACTTCAtCTTTACCCAAAAGAGTGGACTCTGTG A Protein > hT2R54 (SEQ ID NO: 7) MTKLCDPAESELSPFLITLILAV laey IGIIANGFIMAIHAAE VQNK AVSTSGRILVFLSVSRIA QSLMMLEIT SSTSLSFYSEDAVYYAFKISF IF NFCS WFAAWLSFFYFVKIANFSYPLFLK RWRITGLIPWL WLSVF ISFSHS FCINICTVYC NSFPIHSSNST KTYLSEINWGIAFFFNIiGI VTP 1MFILTATL I S KRHT HMGSNATGSNDPS EAHMGAIKAISYF ILYIFNAVALFIYLSN FDINS WNNLCQIIMAAYPASHSIL IQDNPG LRRA KR QLRLHLYPKE TL While the above detailed description has several described embodiments of the present invention, it is to be understood that the foregoing description is illustrative only and not limitative of the disclosed invention. The invention will be limited only by the claims that follow.

Claims (128)

1. CLAIMS 1. An assay for identifying a compound that modulates a human T2R bitter taste receptor that specifically responds to a chlorogenic lactone the hT2R that is selected from the group consisting of hT2R8, hT2R14, and hT2R54, characterized in that it comprises: i. classifying a compound for its effect on a chlorogenic lactone compound to induce the activation of hT2R8, hT2R14, hT2R54 or a fragment, vanant, ortholog, mutant or chimera thereof which is activated by a chlorogenic lactone or a structurally related compound, and 11. to determine whether the compound modulates hT2R8, hT2R14, and / or hT2R54 associated with the bitter taste based on its effect on receptor activation by the chlorogenic lactone or a structurally related compound.
2. 2. The assay according to claim 1, characterized in that the hT2R8 is selected from i. the amino acid sequence contained in SEQ ID NO: 3, 11 is encoded by a nucleic acid sequence that hybridizes under severe hybridization conditions to the sequence contained in SEQ ID NO: 2, or m. it has an amino acid sequence that possesses at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 3.
3. 3. The assay according to claim 1, characterized in that the hT2R14 is selected from i. the amino acid sequence contained in SEQ ID NO: 5, ii. is encoded by a nucleic acid sequence that hybridizes under severe hybridization conditions to the sequence contained in SEQ ID NO: 6, or iii. has an amino acid sequence that possesses at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 5.
4. 4. The assay according to claim 1, characterized in that the hT2R54 is selected from i. the amino acid sequence contained in SEQ ID NO: 7; ii. is encoded by a nucleic acid sequence that hybridizes under severe hybridization conditions to the sequence contained in SEQ ID NO: 6; or iii. it has an amino acid sequence possessing at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 7.
5. 5. The assay according to claim 1, characterized in that the taste receptor is expressed on a cell membrane.
6. 6. The assay according to claim 1, characterized in that the taste receptor is expressed on an isolated cell membrane.
7. 7. The assay according to claim 1, characterized in that the taste receptor is expressed on an intact cell.
8. 8. The assay according to claim 1, characterized in that the taste receptor is expressed on a eukaryotic cell.
9. 9. The assay according to claim 1, characterized in that the taste receptor is expressed by an amphibian, mammalian or insect cell.
10. 10. The assay according to claim 1, characterized in the taste receptor is expressed on a cell selected from a HEK293, BHK, COS, HEK293T, CHO and Xenopus oocyte.
11. 11. The test according to claim 1, characterized in that it is a fluorometric test.
12. 12. The assay according to claim 1, characterized in that it is a binding assay.
13. 13. The test in accordance with the claim 1, characterized in that it detects the effect on the compound when analyzing its effect on an intracellular ion concentration.
14. 14. The assay according to any of claim 1, characterized in that it detects the effect of the compound on intracellular sodium or calcium.
15. 15. The assay according to claim 1, characterized in that it detects the effect of the compound on the cell's central membrane potential.
16. 16. The test in accordance with the claim 1, characterized in that it detects the effect of the compound on the transcription of the taste receptor.
17. 17. The assay according to claim 1, characterized in that the compound is selected based on its ability to block the interaction of the taste receptor with ranitidine.
18. 18. The assay according to claim 1, characterized in that it detects the effect of the compound on intracellular cAMP, cGMP or IP3.
19. 19. The test in accordance with the claim 1, characterized in that the taste receptor comprises the extracellular domain or transmembrane region of the taste receptor.
20. 20. The assay according to claim 1, characterized in that the assay detects changes in calcium using a calcium-specific fluorescent dye.
21. 21. The assay according to claim 1, characterized in that the assay detects changes in intracellular calcium using a dye selected from Fluo-3, Fluo-4 and Fura-2.
22. 22. The assay according to claim 1, characterized in that 1 the receptor is in solution.
23. 23. The assay according to claim 1, characterized in that it is a binding assay that detects changes in spectroscopic characteristics, hydrodynamic characteristics or solubility.
24. 24. The assay according to claim 1, characterized in that it detects the effect of the compound on the taste receptor complex with a protein G.
25. 25. The assay according to claim 1, characterized in that it detects the effect of the compound on the flavor receptor complex with a G protein selected from transducin, gustducin, Gal5, Gai6 or a chimera thereof.
26. 26. The assay according to claim 1, characterized in that it is a fluorescence polarization assay.
27. 27. The assay according to claim 1, characterized in that the taste receptor is bound to a solid phase substrate.
28. 28. The test according to claim 1, characterized in that it is a high performance test.
29. 29. The assay according to claim 1, characterized in that the taste receptor is expressed by a HEK293 cell.
30. 30. An assay for identifying a compound that modulates a bitter taste receptor that specifically responds to 3CoQAL chlorogenic lactone, characterized in that the taste receptor is selected from hT2R8, hT2R14, and hT2R54 comprising: i. classifying a compound for its effect on the structurally related compound 3CoQAL to induce the activation of hT2R8, hT2R14, or hT2R54, or a fragment, variant, ortholog, mutant or chimera thereof that is activated by 3CoQAL or a structurally related compound; and ii. determining whether the compound modulates hT2R8, hT2R14, or hT2R54 based on its effect on receptor activation by 3CoQAL or a structurally related compound.
31. 31. The assay according to claim 30, characterized in that hT2R8 is selected from the group consisting of: i. the polypeptide contained in SEQ ID NO: 3. ii. a polypeptide encoded by a nucleic acid sequence that hybridizes especially to the nucleic acid sequence contained in SEQ ID NO: 2; and iii. a polypeptide having at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 3.
32. 32. The assay according to claim 30, characterized in that hT2R14 is selected from the group consisting of: i. the polypeptide contained in SEQ ID NO: 3. 11. a polypeptide encoded by a nucleic acid sequence that hybridizes specifically to the nucleic acid sequence contained in SEQ ID NO: 2; and m. a polypeptide having at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 3.
33. 33. The assay according to claim 30, characterized in that the hT2R54 is selected from the group consisting of: i. the polypeptide contained in SEQ ID NO: 3. ii. a polypeptide encoded by one by a nucleic acid sequence that hybridizes specifically to the nucleic acid sequence contained in SEQ ID NO: 2; and m. a polypeptide possessing at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 3.
34. 34. The assay according to claim 30, characterized in that the taste receptor is expressed on an isolated cellular membrane.
35. 35. The assay according to claim 30, characterized in that the taste receptor is expressed on an intact cell.
36. 36. The test according to claim 30, 31, characterized in that the taste receptor is expressed on a eukaryotic cell.
37. 37. The assay according to claim 30, characterized in that the taste receptor is expressed by an amphibian, mammalian or insect cell.
38. 38. The assay according to claim 30, characterized in the taste receptor is expressed on a cell selected from a HEK293, BHK, COS, HEK293T, CHO and Xenopus oocyte.
39. 39. The assay according to claim 30, characterized in that it is a fluorometric assay.
40. 40. The test in accordance with the claim 30, characterized in that it is a binding assay.
41. 41. The assay according to claim 30, characterized in that it detects the effect on the compound when analyzing its effect on an intracellular ion concentration.
42. 42. The assay according to claim 30, characterized in that it detects the effect of the compound on intracellular sodium or calcium.
43. 43. The assay according to claim 30, characterized in that it detects the effect of the compound on the cell's central membrane potential.
44. 44. The assay according to claim 30, characterized in that it detects the effect of the compound on the transcription of the taste receptor.
45. 45. The test in accordance with the claim 30, characterized in that the compound is selected based on its ability to block the interaction of the taste receptor with ranitidine.
46. 46. The assay according to claim 30, characterized in that it detects the effect of the compound on intracellular cAMP, cGMP or IP3.
47. 47. The assay according to claim 30, characterized in that the taste receptor comprises the extracellular domain or transmembrane region of the taste receptor.
48. 48. The assay according to claim 30, characterized in that the assay detects changes in calcium using a calcium-specific fluorescent dye.
49. 49. The test in accordance with the claim 30, characterized in that the assay detects changes in intracellular calcium using a dye selected from Fluo-3, Fluo-4 and Fura-2.
50. 50. The test according to claim 30, characterized in that the receiver is in solution.
51. 51. The assay according to claim 30, characterized in that it is a binding assay that detects changes in spectroscopic characteristics, hydrodynamic characteristics or solubility.
52. 52. The test in accordance with the claim 30, characterized in that it is a fluorescence polarization assay.
53. 53. The assay according to claim 30, characterized in that it detects the effect of the compound on the flavor receptor complex with a G protein selected from transducin, gustducin, Ga? 5, Gai6 or a quirtiera thereof.
54. 54. assay according to claim 30, characterized in that it is a fluorescence polarization assay.
55. 55. The assay according to claim 30, characterized in that the taste receptor is bound to a solid phase substrate.
56. 56. The test according to claim 30, characterized in that it is a high performance test.
57. 57. The assay according to claim 30, characterized in that the taste receptor is expressed by a HEK293 cell.
58. 58. An assay for identifying a compound that modulates a receptor a bitter T2R taste receptor that specifically responds to either or both of the chlorogenic lactones 3CQAL and 4CQAL with T2R is selected from the group consisting of hT2R8, hTlR14 and hT2R54, characterized in that it comprises : i. classifying a compound for its effect on the denotonium benzoate or a structurally related compound to induce the activation of hT2R8, hTlR14, and / or hT2R54 or a fragment, variant, ortholog, mutant or chimera thereof which is activated by either or both of 3CQAL and 4CQAL or a structurally related compound; and 11. determining whether the compound modulates hT2R8, hT2R14, and / or hT2R54 based on its effect on receptor activation by either or both of 3CQAL and 4CQAL or a structurally related compound.
59. 59. The test in accordance with the claim 58, characterized in that hT2R8 is selected from the group consisting of: i. the polypeptide contained in SEQ ID NO: 3. n. a polypeptide encoded by a nucleic acid sequence that hybridizes especially to the nucleic acid sequence contained in SEQ ID NO: 2; and m. a polypeptide having at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 3.
60. 60. The assay according to claim 58, characterized in that hT2R14 is selected from the group consisting of: i. the polypeptide contained in SEQ ID NO: 3. n. a polypeptide encoded by a nucleic acid sequence that hybridizes specifically to the nucleic acid sequence contained in SEQ ID NO: 2; and m. a polypeptide that possesses at least 90% sequence identity to the polypeptide contained in SEO ID NO: 3.
61. 61. The assay in accordance with the claim 58, characterized in that the hT2R54 is selected from the group consisting of: i. the polypeptide contained in SEQ ID NO: 3. n. a polypeptide encoded by one by a nucleic acid sequence that hybridizes specifically to the nucleic acid sequence contained in SEQ ID NO: 2; and m. a polypeptide having at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 3.
62. 62. The assay according to the claim 58, characterized in that the taste receptor is expressed on a cell membrane.
63. 63. The assay according to claim 58, characterized in that the taste receptor is expressed on an isolated cell membrane.
64. 64. The assay according to claim 58, characterized in that the taste receptor is expressed on an intact cell.
65. 65. The assay according to claim 58, characterized in that the taste receptor is expressed on a eukaryotic cell.
66. 66. The assay according to claim 58, characterized in that the taste receptor is expressed by an amphibian, mammalian or insect cell.
67. 67. The test in accordance with the claim 58, characterized the taste receptor is expressed on a selected cell of a HEK293, BHK, COS, HEK293T, CHO and Xenopus oocyte.
68. 68. The assay according to claim 58, characterized in that it is a fluorométpco assay.
69. 69. The assay according to claim 581, characterized in that it is a binding assay.
70. 70. The assay according to claim 58, characterized in that it detects the effect on the compound when analyzing its effect on a concentration of intracellular ion.
71. 71. The assay according to any of claim 1, characterized in that it detects the effect of the compound on the sodium or calcium cell.
72. 72. The assay according to claim 58, characterized in that it detects the effect of the compound on the cell's central membrane potential.
73. 73. The assay according to claim 58, characterized in that it detects the effect of the compound on the transcription of the taste receptor.
74. 74. The assay according to claim 58, characterized in that the compound is selected based on its ability to block the interaction of the flavor receptor with denatonium.
75. 75. The test in accordance with the claim 58, characterized in that it detects the effect of the compound on intracellular cAMP, cGMP or IP3.
76. 76. The assay according to claim 58, characterized in that the taste receptor comprises the extracellular domain or transmembrane region of the taste receptor.
77. 77. The assay according to claim 58, characterized in that the assay detects changes in calcium using a calcium-specific fluorescent dye.
78. 78. The assay according to claim 58, characterized in that the assay detects changes in intracellular calcium using a dye selected from Fluo-3, Fluo-4 and Fura-2.
79. 79. The assay according to claim 58, characterized in that 1 the receiver is in solution.
80. 80. The assay according to claim 58, characterized in that it is a binding assay that detects changes in spectroscopic characteristics, hydrodynamic characteristics or solubility.
81. 81. The assay according to claim 58, characterized in that it detects the effect of the compound on the taste receptor complex with a G protein.
82. 82. The assay according to claim 58, characterized in that it detects the effect of the compound on the flavor receptor complex with a G protein selected from transducin, gustducin, Ga? 5, Gai6 or a chimera thereof.
83. 83. The assay according to claim 58, characterized in that it is a fluorescence polarization assay.
84. 84. The assay according to claim 58, characterized in that the taste receptor is bound to a solid phase substrate.
85. 85. The test in accordance with the claim 58, characterized in that it is a high performance test.
86. 86. The assay according to claim 58, characterized in that the taste receptor is expressed by a HEK293 cell.
87. 87. An assay for identifying a compound that modulates a bitter taste receptor that responds specifically to the chlorogenic lactone 4FQAL, characterized in that the taste receptor is selected from hT2R8, hT2R14, and hT2R54 comprising: i. classifying a compound for its effect on the structurally related compound 4 FQAL to induce the activation of hT2R8, hT2R14, or hT2R54, or a fragment, vanant, ortholog, mutant or chimera thereof that is activated by 3CoQAL or a structurally related compound; and 11. determining whether the compound modulates hT2R8, and / or hT2R54 based on its effect on receptor activation by 4 FQAL or a structurally related compound. An assay for identifying a compound that modulates a bitter taste receptor that specifically responds to the 3CoQAL chlorogenic lactone, characterized in that the taste receptor is selected from hT2R8, hT2R14, and hT2R54 comprising:
88. 88. The assay according to claim 87 , characterized in that hT2R8 is selected from the group consisting of: i. the polypeptide contained in SEQ ID NO: 3. n. a polypeptide encoded by a nucleic acid sequence that hybridizes especially to the nucleic acid sequence contained in SEQ ID NO: 2; and iii. a polypeptide having at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 3.
89. 89. The assay according to claim 87, characterized in that hT2R14 is selected from the group consisting of: i. the polypeptide contained in SEQ ID NO: 3. ii. a polypeptide encoded by a nucleic acid sequence that specifically hybridizes to the nucleic acid sequence contained in SEQ ID NO: 2; and iii. a polypeptide having at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 3.
90. 90. The assay according to claim 1, characterized in that the hT2R54 is selected from the group consisting of: i. the polypeptide contained in SEQ ID NO: 3. ii. a polypeptide encoded by one by a nucleic acid sequence that specifically hybridizes to the nucleic acid sequence contained in SEQ ID NO: 2; and iii. a polypeptide having at least 90% sequence identity to the polypeptide contained in SEQ ID NO: 3.
91. 91. The assay according to claim 87, characterized in that the taste receptor is expressed on a cell membrane.
92. 92. The assay according to claim 87, characterized in that the taste receptor is expressed on an isolated cellular membrane.
93. 93. The test in accordance with the claim 87, characterized in that the taste receptor is expressed on an intact cell.
94. 94. The assay according to claim 87, characterized in that the taste receptor is expressed on a eukaryotic cell.
95. 95. The assay according to claim 87, characterized in that the taste receptor is expressed by an amphibian, mammalian or insect cell.
96. 96. The assay according to claim 87, characterized in the taste receptor is expressed on a cell selected from a HEK293, BHK, COS, HEK293T, CHO and Xenopus oocyte.
97. 97. The test according to claim 87, characterized in that it is a fluorometric test.
98. 98. The test in accordance with the claim 87, characterized in that it is a binding assay.
99. 99. The assay according to claim 87, characterized in that it detects the effect on the compound when analyzing its effect on an intracellular ion concentration.
100. 100. The assay according to any of claim 87, characterized in that it detects the effect of the compound on intracellular sodium or calcium.
101. 101. The assay according to claim 87, characterized in that it detects the effect of the compound on the cell's central membrane potential.
102. 102. The assay according to claim 87, characterized in that it detects the effect of the compound on the transcription of the taste receptor.
103. 103. The test in accordance with the claim 87, characterized in that the compound is selected based on its ability to block the interaction of the taste receptor with ranitidine.
104. 104. The assay according to claim 87, characterized in that it detects the effect of the compound on intracellular cAMP, cGMP or IP3.
105. 105. The assay according to claim 87, characterized in that the taste receptor comprises the extracellular domain or transmembrane region of the taste receptor.
106. 106. The assay according to claim 87, characterized in that the assay detects changes in calcium using a calcium-specific fluorescent dye.
107. 107. The assay according to claim 87, characterized in that the assay detects changes in intracellular calcium using a dye selected from Fluo-3, Fluo-4 and Fura-2.
108. 108. The assay according to claim 87, characterized in that 1 the receiver is in solution.
109. 109. The assay according to claim 871, characterized in that it is a binding assay that detects changes in spectroscopic characteristics, hydrodynamic characteristics or solubility.
110. 110. The test in accordance with the claim 87, characterized in that it detects the effect of the compound on the flavor receptor complex with a G protein.
111. 111. The assay according to claim 87, characterized in that it detects the effect of the compound on the flavor receptor complex with a G protein. selected transducina, gustducina, Ga? 5, Ga? eo a chimera of the same.
112. 112. The assay according to claim 87, characterized in that it is a fluorescence polarization assay.
113. 113. The assay according to claim 87, characterized in that the flavor receptor is bound to a solid phase substrate.
114. 114. The test according to claim 87, characterized in that it is a high performance test.
115. 115. The assay according to claim 87, characterized in that the taste receptor is expressed by a HEK293 cell.
116. 116. A method for lightening the bitter taste of a food, beverage or medicinal substance, characterized in that it comprises adding thereto an amount that inhibits the bitter taste of a compound identified using any test according to claim 1.
117. 117. A method for lightening the bitter taste of a food, drink or medicinal substance, characterized in that it comprises adding thereto an amount that inhibits the bitter taste of a compound identified using any test according to claim 30.
118. 118. A method to lighten the bitter taste of a food, drink or medicinal substance, characterized in that it comprises adding thereto an amount that inhibits the bitter taste of a compound identified using any test according to claim 58.
119. 119. A method for lightening the bitter taste of a food, drink or medicinal substance, characterized in that it comprises adding to it a quantity ad that inhibits the bitter taste of an identified compound using any test according to claim 87.
120. 120. The method according to claim 116 characterized in that the food, beverage or medicinal substance comprises at least one chlorogenic lactone compound that contributes to its bitter taste.
121. 121. The method according to claim 20, characterized in that the food, beverage OR medicinal substance is a coffee beverage or coffee flavor food or medicinal agent.
122. 122. The method according to claim 120, characterized in that the chlorogenic lactone is selected from the group consisting of 3CoQAL, 4CQAL, 4CQAL and 4 FQAL.
123. 123. A method for lightening the bitter taste of a food, beverage or medicinal substance, characterized in that it comprises removing at least one compound identified in any of the assays of claim 1 that induces activation or increases activation of hT2R8 , hT2R14 and / or hT2R54 by a chlorogenic lactone compound.
124. 124. The method of compliance with the claim 123, characterized in that the compound is a chlorogenic lactone.
125. 125. The method of compliance with the claim 124, characterized in that the chlorogenic lactone is selected from 3CoQAL, 3CQAL, 4CQAL and 4 FQAL.
126. 126. The method according to claim 123, characterized in that the removal is from a coffee beverage or coffee flavor food or medicinal substance.
127. 127. The method according to claim 126, characterized in that toasting or processing a coffee material under conditions that reduce the amount of at least one chlorogenic lactone that contributes to the bitter taste.
128. 128. A method for identifying a food, beverage or medicinal substance that reduces bitterness compared to other foods, beverages or medicinal substances, respectively, characterized in that it comprises a. prepare extracts from a set of foods, beverages or medicinal substances; b. contacting an aliquot of the extract using an assay described in claim 1; c. record the test response with the various aliquots; d. classify the responses of the different extracts; and e. select the sample of food, drink or medicinal substance with the lowest activation activity.