Identifying Umami Peptides Specific to the T1R1/T1R3 Receptor via Phage Display.

Umami peptides are small molecular weight oligopeptides that play a role in umami taste attributes. However, the identification of umami peptides is easily limited by environmental conditions, and the abundant source and high chromatographic separation efficiency remain difficult. Herein, we report a robust strategy based on a phage random linear heptapeptide library that targets the T1R1-Venus flytrap domain (T1R1-VFT). Two candidate peptides (MTLERPW and MNLHLSF) were readily identified with high affinity for T1R1-VFT binding (KD of MW-7 and MF-7 were 790 and 630 nM, respectively). The two peptides exhibited umami taste and significantly enhanced the umami intensity when added to the monosodium glutamate solution. Overall, this strategy shows that umami peptides could be developed via phage display technology for the first time. The phage display platform has a promising application to discover other taste peptides with affinity for taste receptors of interest and has more room for improvement in the future.

Umami taste evaluation based on a novel mouse taste receptor cell-based biosensor.

Umami, a taste sensation known for its savory and delicious properties, has garnered considerable attention from both consumers and the food industry. However, current understanding and evaluation of umami characteristics remain limited, presenting a long-standing issue. To address this challenge, we have developed a self-assembled biosensor based on matured taste receptor cells (TRCs), obtained through isolation and culture of taste stem cells. TRCs, as the recognition element, were mounted onto the surface of a glassy carbon electrode (GCE) treated with gold nanoparticles (AuNPs) and poly-L-lysine (PLL). Key parameters including the cell incubation time and concentration were optimized to ensure the optimal performance of the TRCs-based biosensor. AuNPs were deposited onto the GCE surface via 90 s electrochemical reduction. TRCs concentration of 106 cells/mL and incubation time of 12 h were chosen by electrochemical characterization. Using this novel, rapid, and sensitive TRCs-based biosensor, we successfully detected L-monosodium glutamate (MSG) and other umami substances, demonstrating a good linear relationship within the range of 10-9 - 10-5 M between response signals and concentration of MSG stimuli. Our results provide insights into taste signal transduction mechanisms and suggest the potential for biomimetic sensors in intelligent perception applications.

Bimetallic bionic taste sensor for perception of the synergistic effect of umami substances.

Synergistic effect is one of the main properties of umami substances, elucidating the synergistic effect of umami is of great significance in the food industry. In this study, a bimetallic bionic taste sensor was developed to evaluate the synergistic effect of umami substances based on the perceptual mechanism of the human taste system. The Venus flytrap domain of T1R1 which is in charge of recognizing umami ligands was employed as the sensing element and self-assembled on the bimetallic nanomaterial (MoS2-PtPd) by Au-S bonding, the binding of receptors and ligands is characterized by changes of electrical signals. The sensor had good linearity (R2 > 0.99) and wide detection range in the detection of different kinds of umami substances (amino acids, nucleotides, organic acids, umami peptides) with detection limits as low as 0.03 pM. Comparing with electronic tongues, the sensor owned multiple characteristics of human taste system and could recognize the presence of synergistic effect of umami substances in a variety of real samples. Moreover, the differences in synergistic effect at different concentrations and ratios were also explored, the findings showed that the synergistic effect was more obvious at lower concentrations and balanced ratios of multiple umami substances added. The strategy would afford a promising platform for in-depth research on the mechanism of synergistic effect and multifunctional industrial applications.

Conserved Sites and Recognition Mechanisms of T1R1 and T2R14 Receptors Revealed by Ensemble Docking and Molecular Descriptors and Fingerprints Combined with Machine Learning.

Taste peptides, as an important component of protein-rich foodstuffs, potentiate the nutrition and taste of food. Thereinto, umami- and bitter-taste peptides have been ex tensively reported, while their taste mechanisms remain unclear. Meanwhile, the identification of taste peptides is still a time-consuming and costly task. In this study, 489 peptides with umami/bitter taste from TPDB (http://tastepeptides-meta.com/) were collected and used to train the classification models based on docking analysis, molecular descriptors (MDs), and molecular fingerprints (FPs). A consensus model, taste peptide docking machine (TPDM), was generated based on five learning algorithms (linear regression, random forest, gaussian naive bayes, gradient boosting tree, and stochastic gradient descent) and four molecular representation schemes. Model interpretive analysis showed that MDs (VSA_EState, MinEstateIndex, MolLogP) and FPs (598, 322, 952) had the greatest impact on the umami/bitter prediction of peptides. Based on the consensus docking results, we obtained the key recognition modes of umami/bitter receptors (T1Rs/T2Rs): (1) residues 107S-109S, 148S-154T, 247F-249A mainly form hydrogen bonding contacts and (2) residues 153A-158L, 163L, 181Q, 218D, 247F-249A in T1R1 and 56D, 106P, 107V, 152V-156F, 173K-180F in T2R14 constituted their hydrogen bond pockets. The model is available at http://www.tastepeptides-meta.com/yyds.

Typical Umami Ligand-Induced Binding Interaction and Conformational Change of T1R1-VFT.

Umami taste receptor type 1 member 1/3 (T1R1/T1R3) heterodimer has multiple ligand-binding sites, most of which are located in T1R1-Venus flytrap domain (T1R1-VFT). However, the critical binding process of T1R1-VFT/umami ligands remains largely unknown. Herein, T1R1-VFT was prepared with a sufficient amount and functional activity, and its binding characteristics with typical umami molecules (monosodium l-glutamate, disodium succinate, beefy meaty peptide, and inosine-5'-monophosphate) were explored via multispectroscopic techniques and molecular dynamics simulation. The results showed that, driven mainly by hydrogen bond, van der Waals forces, and electrostatic interactions, T1R1-VFT bound to umami compound at 1:1 (stoichiometric interaction) and formed T1R1-VFT/ligand complex (static fluorescence quenching) with a weak binding affinity (Ka values: 252 ± 19 to 1169 ± 112 M-1). The binding process was spontaneous and exothermic (ΔG, -17.72 to -14.26 kJ mol-1; ΔH, -23.86 to -12.11 kJ mol-1) and induced conformational changes of T1R1-VFT, which was mainly reflected in slight unfolding of α-helix (Δα-helix < 0) and polypeptide chain backbone structure. Meanwhile, the binding of the four ligands stabilized the active conformation of the T1R1-VFT pocket. This work provides insight into the binding interaction between T1R1-VFT/umami ligands and improves understanding of how umami receptor recognizes specific ligand molecules.

A novel umami electrochemical biosensor based on AuNPs@ZIF-8/Ti3C2 MXene immobilized T1R1-VFT.

The bioelectronic tongues based on taste receptors have been emerging with human-like taste perception. However, the practical applications of the receptor-based biosensors were restricted by their narrow and low dynamic ranges. Here, a novel immobilization strategy based on AuNPs@ZIF-8/Ti3C2 MXene was developed to immobilize the umami ligand binding domain (T1R1-VFT), to fabricate an umami biosensor for umami substances detection. Through the synergic effect of AuNPs@ZIF-8 and Ti3C2 MXene, the capacity to load T1R1-VFT was effectively increased, and the response signal was also amplified by approximately 3 times. The proposed biosensor showed an ultrawide dynamic range of 10-11-10-3 M, and a high upper limit of detection, which was closer to the human taste threshold and suitable for detecting foods rich in umami substances. Additionally, the biosensor was successfully applied to detect real samples and analyze the synergistic effects of binary umami substances.

Biomimetic ion nanochannels for sensing umami substances.

Umami is one of the basic taste sensing, and represents the recognition of N-containing compounds capable of evaluating the nutritious contents of food. Although several sensors have been developed, the assessment of umami intensity remains challenging due to the limitations of sensor specificity, sensitivity, and performance stability. Here we present a biomimetic conical nanochannels system integrated with Venus flytrap (VFT) domain from human umami receptor T1R1 subunit to meet the concern. By taking advantage of sensitive transmembrane ionic flux change, the functional nanochannels could precisely distinguish umami substances from other tastants. Detailed mechanism analysis reveals that specific binding between T1R1 and umami substances triggers local conformation change and surface charge redistribution of the protein, which modulates the ionic current. This study initiates the application of nanochannel device in taste perception, which could help to disclose umami perception mechanism and screen new umami substances.

Taste and stability characteristics of two key umami peptides from pufferfish (Takifugu obscurus).

Takifugu obscurus (T. obscurus) is known for its umami taste. Two taste-active peptides, Pro-Val-Ala-Arg-Met-Cys-Arg (PR-7) and Tyr-Gly-Gly-Thr-Pro-Pro-Phe-Val (YV-8), were proved as key compounds that contributed to the typical taste of T. obscurus. However, whether these peptides have the potential as umami supplements is unknown. The purpose of this study was to investigate the taste characteristics of PR-7 and YV-8, as well as stability at different pH values by sensory evaluation, instrumental analysis and quantum chemical calculation. The results indicated that PR-7 and YV-8 presented umami taste at near neutral pH (6.5-8.0) and had umami-enhancing effects. PR-7 also exhibited significant kokumi activity. Additionally, two peptides showed remarkable stability after different pH treatments, especially YV-8; this may be related to its stable structural property. All the results suggest that both peptides have great potential to be applied in complex foods to provide desirable taste, and act as a feasible alternative to monosodium l-glutamate.

Study on the distribution of umami receptors on the tongue and its signal coding logic based on taste bud biosensor.

Taste signals are uniformly encoded and transmitted to the brain's taste center by taste buds, and the process has not been systematically studied for several decades. The aim of this work was to investigate the distribution of umami receptors on the tongue and its signal coding logic based on the taste bud biosensors. Taste bud biosensors were constructed by immobilizing the taste bud tissues from different tongue regions of the rabbit to the glassy carbon electrode surface; The Shennong information equations were used to analysis the pattern of umami receptors to encode ligands information; The signal amplification capabilities of two types umami receptors (T1R1/T1R3 and mGluRs) were analyzed for the two ligands (L-monosodium glutamate (MSG) and disodium 5'-inosinate (IMP)). The results showed that each taste bud biosensor could sense MSG and IMP with different response currents based on enzyme-substrate kinetics. There was only a small fraction of a great quantity of metabotropic glutamate receptors (mGluRs) could be activated to encode MSG signal. Importantly, T1R1 was more expressed in the rostral tongue cells whose sensitivity to MSG was nearly 100 times stronger than that of caudal tongue cells. The method we proposed made it possible to reveal the distribution and signals coding logic of umami receptors for ligands, which showed great potential to explain the interaction mechanism of umami substances with their receptors more accurately and to develop of artificial intelligent taste sensory.

Human-like performance umami electrochemical biosensor by utilizing co-electrodeposition of ligand binding domain T1R1-VFT and Prussian blue.

Over the past decades, due to the desire for artificial umami flavors, apparatuses for detecting the umami taste have constantly been developed. Nevertheless, most information on umami is still acquired through human sensory assessment, which makes it difficult to establish an umami standard or quantify the umami flavor. In this study, the ligand binding domain called venus flytrap (VFT) domain of the umami taste receptor protein T1R1 was used as a recognition element, and an electrochemical biosensor based on a double-signal amplification strategy was constructed using single-walled carbon nanotubes (SWCNTs) and Prussian blue (PB). Moreover, the umami taste of four representative umami substances, inosine-5'-monophosphate (IMP), monosodium L-glutamate (MSG), beefy meaty peptide (BMP), and sodium succinate (WSA), were successfully quantitatively measured using differential pulse voltammetry (DPV) at an electrochemical workstation. Based on an equation (S/N = 3), the low detection limits (LODs) of IMP, MSG, BMP, and WSA were 0.1, 0.1, 0.1, and 0.01 pM, respectively. Meanwhile, a normalized signal intensity of more than 90% was kept for 4 days. The results showed that the biosensor could be used to detect umami substances with high sensitivity and selectivity, and was shown to have human-like performance. To develop the T1R1-VFT biosensor using the above-mentioned method, we utilized the ligand binding domain of the human umami receptor, rather than the entire umami receptor protein, which had a complex structure, having the following advantages: volume reduction, simplicity, and stability. This method has great potential for the detection of umami tastes, instead of using sensory evaluation, and for the development of new artificial flavorings.

Sensory-Guided Analysis of Key Taste-Active Compounds in Pufferfish (Takifugu obscurus).

To investigate key taste-active components in Takifugu obscurus, 28 putative taste compounds in cooked muscle of T. obscurus were quantitatively analyzed and the pivotal components were identified by taste reconstitution, omission, and addition tests. Moreover, the role of flavor peptides in the overall taste profile of T. obscurus was evaluated. Sensory evaluation revealed that glutamic acid, serine, proline, arginine, lysine, adenosine 5'-monophosphate, inosine 5'-monophosphate (IMP), succinic acid, sodium, potassium, phosphates, and chlorides were the core taste-active contributors to T. obscurus. Besides glutamic acid, IMP, succinic acid, and potassium, the characteristic T. obscurus-like umami and kokumi profiles were induced by adding flavor peptides, among which Pro-Val-Ala-Arg-Met-Cys-Arg and Tyr-Gly-Gly-Thr-Pro-Pro-Phe-Val were identified as key substances on the basis of the addition test and dose-response analysis. The present data may help to reveal the secret of the delicious taste of T. obscurus and provide the basis for the development of deeper flavor analysis of pufferfish.