Screening method and application of bifunctional peanut peptide with salty taste enhancing and antioxidant activity and product

Abstract

The invention discloses a salty taste enhancing peptide screened from peanut protein and a screening method of the salty taste enhancing peptide, and belongs to the technical field of food seasonings. Four peptide fragments SPDIY, DPSPR, QPGDY and SPPGER with a potential salty taste enhancing effect are obtained through screening according to a molecular docking result, and the molecular weights of the four peptide fragments are all smaller than 1000 Da. Sensory evaluation and electronic tongue analysis show that the four peptide fragments all have a good salty taste enhancing effect and can be applied to the field of food seasonings, and the purpose of reducing salt but not reducing salty taste is achieved. Wherein the peptide fragment of which the amino acid sequence is QPGDY is a bifunctional active peptide, and the peptide fragment has in-vitro antioxidant activity while reducing salt. The invention provides a theoretical basis for finding new salty taste enhancing peptide resources in peanuts and developing nutritional and healthy salty taste enhancing peptides.

Description

Technical Field

[0001] The invention relates to the technical field of food seasonings, and in particular to a salty taste enhancing peptide screened from peanut protein and capable of docking with a salty taste receptor TMC4 and an application thereof, and the peptide also has antioxidant activity. Background Art

[0002] Table salt (NaCl) is an important component of saltiness in our daily diet and plays an important role in our daily lives. From a food perspective, saltiness usually comes from sodium ions, which are obtained through salt or other sodium-containing compounds. The World Health Organization's goal is to reduce the global average daily salt intake by 30% by 2025. Therefore, how to "reduce salt without reducing saltiness" without affecting the sensory quality of food has attracted widespread attention from all over the world.

[0003] Salty taste enhancing peptides have attracted much attention as a natural, safe, accessible, low taste threshold, and nutritious salt substitute. In recent years, computer methods have been widely used in peptide screening and mechanism studies. Compared with traditional methods, these methods can quickly and accurately identify the best bioactive compounds by simulating and predicting the interactions of biomacromolecules, which may solve time and cost issues. Therefore, this study used a combination of computer technology and sensory evaluation to quickly screen salty taste enhancing peptides. Transmembrane channel-like 4 (TMC4) is a voltage-dependent chloride ion channel that is mainly expressed in the posterior part of the taste buds and is responsible for salt perception. It has been shown to be involved in the perception of high concentrations of salty taste through mouse experiments and exhibits a high salt effect at low salt concentrations. As a new salt taste receptor, TMC4 can reveal different signal transduction pathways and help to gain a deeper understanding of the diversity, complexity, and novelty of salt taste perception. Therefore, as a new type of specific salt receptor, TMC4 provides a new way to explore taste enhancement. Peanut protein is an important plant protein that is widely used in food and nutritional supplements due to its high nutritional value and strong functionality. However, there is no clear classification boundary for peanut protein at present. This study mainly adopted the systematic allergen nomenclature method and focused on the main seed storage proteins in peanuts: Arah1, Ara h2, Arah3 and Ara h6 accounted for 57%-65% of the total peanut protein content. In addition, we also included conarachidin A and conarachidin B in our study. Therefore, we used peanut protein as raw material and adopted high-throughput virtual screening technology to quickly screen potential salty taste-enhancing peptides.

[0004] Based on this, the present invention aims to screen out novel salty taste enhancing peptides from peanut protein using virtual screening technology, providing a new strategy for achieving “salt reduction” without “salt reduction”. Summary of the invention

[0005] In view of this, the present invention provides a method for screening peanut peptides with both salty taste enhancement and anti-oxidation properties, applications and products.

[0006] In order to achieve the above object, the present invention adopts the following technical solution:

[0007] The present invention provides a salty taste enhancing peptide which also has an antioxidant effect. The peptide is screened from peanut protein and binds to TMC4 receptor protein. The peptide is composed of one of the following:

[0008] The amino acid sequence is SPDIY, and the molecular weight is 593.63Da;

[0009] The amino acid sequence is DPSPR, and the molecular weight is 570.6Da;

[0010] The amino acid sequence is QPGDY, and the molecular weight is 578.57 Da;

[0011] The amino acid sequence is SPPGER and the molecular weight is 641.67Da.

[0012] Furthermore, the peptides with the amino acid sequences of SPDIY, QPGDY and DPSPR can be docked with the TMC4 receptor based on molecular docking, and the main binding sites are Lys136, Arg140, Gln143, Ala158, Glu161, Ser162, Ser165, Arg168, Leu268, Arg272, Ser275 and Gln279. Among them, Arg272, Glu161 and Arg168 have the highest frequency of occurrence and are key amino acid residues;

[0013] Furthermore, the peptide segment with the amino acid sequence of SPPGER can be docked with the TMC4 receptor based on molecular docking, and the main binding sites are Gln143, Glu161, Ser162, Arg168, Ser165, Arg272 and Gln279. Among them, Ser165 and Gln279 have the highest frequency of occurrence and are key amino acid residues.

[0014] Furthermore, the saltiness value of the peptide segment with the amino acid sequence of SPDIY is 4.04, the saltiness enhancement threshold is 0.64 mmol / L, and it can replace about 6.32% of NaCl;

[0015] and / or, the peptide segment with the amino acid sequence of DPSPR has a saltiness value of 7.1, a saltiness enhancement threshold of 0.16 mmol / L, and can replace about 87.1% of NaCl;

[0016] And / or, the peptide segment with the amino acid sequence of QPGDY has a saltiness value of 4.42, a saltiness enhancement threshold of 0.33 mmol / L, and can replace about 16.3% NaCl;

[0017] And / or, the peptide segment with the amino acid sequence of SPPGER has a saltiness value of 4.49 and a saltiness enhancement threshold of 0.30 mmol / L, and can replace about 18.2% NaCl.

[0018] Furthermore, the peptide segment with the amino acid sequence of QPGDY can scavenge at least one of DPPH free radicals and ABTS free radicals and has antioxidant activity.

[0019] The present invention provides a method for screening salty taste enhancing peptides, characterized in that the steps are as follows:

[0020] (1) Construct a TMC4 salty taste receptor protein model using the AlphFold2 online server and evaluate its reliability;

[0021] The reliability evaluation: pLDDT score>70; the proportion of amino acid residues falling in the allowed region and the maximum allowed region in the whole protein is required to be higher than 90%, and the conformation of the model is considered reasonable and reliable; according to the Ramachandran diagram, the proportion of amino acid residues falling in the allowed region and the maximum allowed region in the whole protein is required to be higher than 90%;

[0022] (2) Computer simulation of enzymatic hydrolysis of peanut protein to obtain enzymatic peptides with a sequence length of 4-8;

[0023] (3) predicting the biological activity of the enzymatic peptides, screening out peptides with potential biological activity, predicting their physicochemical properties, and screening out non-toxic and water-soluble peptides;

[0024] Among them, the screening criteria for the biological activity prediction are: prediction score>0.5;

[0025] The screening criteria for toxicity prediction are: NonToxin;

[0026] The screening criteria for water solubility prediction are: Good water solubility;

[0027] (4) molecular docking the TMC4 receptor protein model with the non-toxic, water-soluble peptide segment of step (3) to form a small molecule ligand and receptor protein complex system;

[0028] Among them, the TMC4 receptor protein needs to remove water molecules and add hydrogen atoms in the CHARMM force field environment;

[0029] The active site coordinates of the molecular docking are: X=-15.25, Y=11, Z=7.25, and the radius is 30;

[0030] The molecular docking site coordinates select the largest cavity in the TMC4 receptor protein as the docking pocket;

[0031] (5) Screening peptides with potential salt-increasing effects based on the molecular docking results and synthesizing the peptides by solid-phase synthesis;

[0032] Wherein, the screening: selecting peptides with low energy values ​​in CDOCKER energy;

[0033] (6) The synthesized peptides are subjected to sensory evaluation and electronic tongue analysis to obtain the above-mentioned salty taste enhancing peptides.

[0034] The invention provides an application of a salty taste enhancing peptide in the preparation of condiments and foods.

[0035] The invention provides a product containing the salty taste enhancing peptide screened from peanut protein.

[0036] Compared with the prior art, the beneficial effects achieved by the present invention are as follows: the present invention screened out four peptides, SPDIY, DPSPR, QPGDY and SPPGER, with a molecular weight of less than 1000Da from peanut protein, and sensory evaluation and electronic tongue analysis showed that these four peptides all have a good salt-enhancing effect. These four salty taste enhancing peptides can be applied to food seasonings to achieve the purpose of "reducing salt" without "reducing saltiness". Among them, the peptide with the amino acid sequence of QPGDY is a bifunctional active peptide, which has in vitro antioxidant activity while reducing salt. The new peptide sequence obtained by the present invention provides a new resource for the development of functional foods. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on the provided drawings without paying creative work.

[0038] Figure 1 The evaluation diagram of the TMC4 salty taste receptor protein model of the embodiment of the present invention, wherein (a) is the TMC4 receptor protein model, (b) is the pLDDT score of the modeling result, (c) is the Ramachandran diagram, and (d) is the Verify3D diagram;

[0039] Figure 2 This is a diagram showing the results of implementing the electronic tongue of the present invention;

[0040] Figure 3 This is a result diagram of the salinity value analysis of an embodiment of the present invention;

[0041] Figure 4 The statistical results of the amino acid sites of the peptide segments of the present invention and the TMC4 receptor protein docking;

[0042] Figure 5 The main interaction forces and occurrence frequencies of the salty taste enhancing peptides of the present invention and TMC4;

[0043] Figure 6 The 2D and 3D interaction diagrams of the four peptide segments of the present invention and the TMC4 salty taste receptor protein molecule are shown;

[0044] Figure 7 is the DPPH free radical scavenging rate of the four peptides in Examples of the present invention;

[0045] Figure 8 ABTS free radical scavenging rates of the four peptides in Examples of the present invention. DETAILED DESCRIPTION

[0046] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0047] The drugs required for the present invention are conventional experimental drugs, which are purchased from commercial channels; the experimental methods not mentioned are conventional experimental methods and will not be described in detail here.

[0048] Example 1

[0049] Construction of TMC4 receptor protein model

[0050] The amino acid sequence of TMC4 (NP_001138775) was obtained from the NCBI database, and the NCBIBLAST tool was used to search for the amino acid sequence of TMC4 with a homology of less than 30%. Therefore, the AlphFold2 online server was used to model the TMC4 amino acid using the AI ​​de novo folding algorithm, and the TMC4 salty taste receptor protein model was obtained. The final constructed 3D receptor protein model is shown in Figure 1 (a).

[0051] Reliability evaluation of TMC4 receptor protein model

[0052] The reliability of the TMC4 receptor protein model was evaluated based on the pLDDT score. A pLDDT score of ≥ 90 indicated that the model had a high degree of confidence, and a pLDDT score between 70 and 90 indicated that the model had a good degree of confidence. Figure 1 As shown in (b), the pLDDT values ​​of the five top-ranked protein models (rank_1, rank_2, rank_3, rank_4 and rank_5) are 78.9, 77.1, 76.2, 74.3 and 72, respectively. Among them, rank_1 has the highest pLDDT value (78.9), so rank_1 is selected as the preliminary experimental model for the next step of evaluation. The rank_1 model is evaluated using the residual percentage of the Ramachandran plot and ERRAT in SEVESv6.0. The evaluation results are shown in Figure 1 As shown in (c), about 92.7% of the amino acid residues are located in the largest allowed region, 6.7% of the amino acid residues are located in other allowed regions, only 0.2% of the amino acid residues are located in the disallowed region, and 0.5% of the residues are located in the prohibited region. Therefore, the TMC4 receptor constructed by the present invention is reasonably structured, indicating that the model constructed by the AI ​​de novo folding algorithm has high reliability and can be used for subsequent research. Finally, the overall quality of the protein crystal structure was evaluated using the ERRAT method ( Figure 1 d), the Overall quality factor was 98.102%, and the model was considered reliable. In conclusion, these results validated the reliability of the TMC4 protein model as a salty taste receptor.

[0053] Computer simulation of enzymatic hydrolysis of peanut protein

[0054] The protein subunits of Ara h1, Ara h2, Ara h3, Ara h6, conarachidin A and conarachidin B were obtained from the NCBI GenBank database, with accession numbers AAL27476.1, QGA89299.1, ACH91862.1, AAM78596.1, AEL30377.1 and AAU21493.1, respectively. The protein sequences obtained above were simulated enzymatically digested using pepsin (EC3.4.23.1), trypsin (EC3.4.21.4) and chymotrypsin (EC3,4,21,1) on the ExPASyPeptideCutter online website, and a total of 307 peptides were obtained. Generally, good biological activity is the key to studying peptides. The biological activity of each peptide was predicted using the Peptide Ranker website. After entering the amino acid sequence of each peptide, there will be a corresponding biological activity score. The higher the score, the greater the biological activity of the peptide. Usually, a peptide with a biological activity score of not less than 0.5 can be considered an active peptide. In addition, non-toxicity is a basic requirement for any potential biomacromolecule, and good water solubility helps to improve the bioavailability of peptides. The online website Innovagen was used to analyze the solubility of peptides in water, and the website divided the water solubility of peptides into two types: "Good water solubility" and "Poor water solubility". The online website Toxin Pred was used to predict the toxicity of active peptides. The final results are shown in Table 1. A total of 19 peptides with high biological activity, non-toxicity and good water solubility were enzymatically hydrolyzed. By cross-referencing the reported amino acid sequences in the database and literature, we avoided the similarity and redundancy of short peptide sequences, while also taking into account the complexity, time and cost of synthesizing long peptide chains. Therefore, we selected peptides of 4 to 8 amino acids as the research focus. A total of 13 potential salty taste-enhancing peptides were predicted, including tripeptides QPDY, QPPR and GQGR; pentapeptides QPDGY, SPDIY, DPSPR, GPSPR and ERPTF; hexapeptides GPAGPR, SPPER and GRPCVM; and heptapeptide QSQRPPR.

[0055] Table 1 Prediction of biological activity and analysis of physicochemical properties of peptides

[0056]

[0057] Screening of potential salty taste enhancing peptides

[0058] In order to gain a deeper understanding of the interaction mechanism between peptides and the TMC4 salty taste receptor protein model, the TMC4 receptor was added to the DS software, and the solvent water molecules were removed and hydrogen atoms were added under the CHARMM force field environment. The constructed peptide was docked into the active cavity of the TMC4 structure using CDOCKER of the Discovery Studio software. The docking active position was: X = -15.25, Y = 11, Z = 7.25, the radius was 30, and the other parameters were the default values. The CDOCKER molecular docking results are shown in Table 2. All 13 potential salt-increasing peptides can enter the docking area of ​​the TMC4 receptor and show strong interactions. This may be because the binding region of TMC4 has an open conformation and a large binding cavity. It is worth noting that the docking energy values ​​of 4-5 amino acids with TMC4 ranged from -45.73 kcal / mol to -89.2205 kcal / mol, while the docking energy values ​​of 6-7 amino acids ranged from -53.2087 kcal / mol to -79.5969 kcal / mol. The level of docking energy usually reflects the stability of the ligand-receptor complex binding. Lower docking energy indicates more stable binding and a higher probability of potential activity. Therefore, based on the docking energy analysis results of the 13 peptides, we selected the top four peptides with the lowest docking energy, QPGDY, SPDIY, DPSPR, and SPPGER, for further study. In addition, these peptides have not been recorded in the literature, indicating that they may be undiscovered salty taste enhancing peptides. Subsequently, these peptides were synthesized and characterized to study their taste characteristics.

[0059] Table 2 Molecular docking results of potential salty miso peptides and TMC4 receptor protein

[0060]

[0061] Solid phase peptide synthesis

[0062] Nanjing Peptide Valley Biotechnology Co., Ltd. was commissioned to synthesize peptides through solid phase synthesis.

[0063] Taste characteristics of synthetic peptides and electronic tongue analysis

[0064] To determine the saltiness intensity of synthetic peptides, a sensory evaluation panel used the taste dilution method to measure the saltiness threshold and then conducted taste descriptions. In ultrapure water, the four synthetic peptides only showed weak tastes and no detectable saltiness. From this, we can conclude that these four peptides do not have saltiness by themselves. However, it is worth noting that when mixed with NaCl solution, they all enhance the saltiness of the mixed solution. Specifically, QPGDY shows saltiness and weak sweetness, SPDIY shows saltiness, weak umami and weak sweetness, DPSPR shows saltiness and weak umami, and SPPGER shows saltiness, sweetness and weak umami. The ranking of the saltiness enhancement thresholds is as follows: DPSPR (0.16 mmol / L) < SPPGER (0.30 mmol / L) < QPGDY (0.33 mmol / L) < SPDIY (0.64 mmol / L). Using a 0.35% NaCl solution as a positive control, the saltiness enhancement effects of the four peptides were analyzed using an electronic tongue. As Figure 2 shown, the overall tastes of the four saltiness-enhancing peptides showed a similar trend, with saltiness being the main taste affecting the overall taste, followed by umami, and sourness being the weakest. The significant differences in acidity values may be related to the solubility and amino acid composition of the peptides. Research has shown that the solubility of peptides affects their dissociation behavior in water, leading to an increase in acidity. In addition, the addition of acidic residues and acetate during the polypeptide synthesis process is another key factor contributing to the increase in acidity values. In terms of the saltiness enhancement effect, DPSPR had the most significant saltiness enhancement effect, with the salt value increasing by 87.1%, followed by SPPGER (18.2%), QPGDY (16.3%) and SPDIY (6.32%).

[0065] Sensory evaluation

[0066] The sensory evaluation panel consisted of 12 laboratory members, including 8 females and 4 males, aged between 20-30 years old, in good physical condition, with good taste and no bad habits. They were trained in sensory training according to ISO 8586-1:2012 to enable them to judge the concentration of five basic flavor solutions. The flavor reference solutions were prepared as follows: 0.08% quinine sulfate solution (bitter), 0.08% citric acid solution (sour), 0.35% sodium chloride solution (salty), 0.35% monosodium glutamate solution (fresh), 1.00% sucrose solution (sweet). The sensory evaluation panel members performed sensory evaluations on a scale of 0-6 points, with 0 representing no taste and 6 representing the strongest taste. Five reference solutions of different flavor intensity were prepared, among which the salty sensory evaluation reference solution was 0.35% NaCl solution, with a salty score of 3 points, and the salty score increased or decreased by 1 point for every 0.1% increase or decrease in concentration. The panel members were asked to rinse their mouths with 30-40 ml of ultrapure water for 5 minutes between testing two different samples. The synthetic peptides were dissolved in 0.4% NaCl solution to prepare a peptide solution with a concentration of 0.02%. Then different panelists scored and recorded the five flavors of the peptide solution in turn. The results showed that all four peptides had a salt-enhancing effect. The sensory evaluation results were as follows: Figure 3 As shown. Compared with the control group, the four peptide solutions with a concentration of 0.02% showed stronger salty characteristics. Among them, the peptide DPSPR had the highest salty intensity (4.7), which was roughly equivalent to the salty intensity of 0.45% NaCl solution. These results are basically consistent with the electronic tongue. In summary, the development of a new flavor substance by combining salty taste enhancing peptides with NaCl can effectively reduce the intake of Na ions without reducing the salt intensity.

[0067] Molecular mechanism of salty taste enhancing peptides

[0068] The four salty taste-enhancing peptides were subjected to multiple molecular docking experiments with TMC4 to determine the best binding posture for each ligand-receptor docking. The molecular docking energy values ​​showed that the four salty taste-enhancing peptides could enter the active cavity of TMC4 with lower energy. The average molecular docking energies of the pentapeptide and hexapeptide were –84.4025 kcal / mol and –79.5969 kcal / mol, respectively, indicating that the hexapeptide was easier to bind to TMC4 than the pentapeptide. However, although SPDIY had the lowest docking energy with TMC4, sensory evaluation and electronic tongue analysis showed that DPSPR had the most significant salt-enhancing effect. This shows that the docking energy value cannot be used alone as the sole criterion for evaluating the intensity of salty taste. In order to further clarify the interaction between the four peptides and the TMC4 receptor protein, we analyzed the binding sites of the salty taste-enhancing peptides with TMC4. Figure 4As shown, the binding sites of the pentapeptide (SPDIY, QPGDY and DPSPR) with TMC4 include Lys136, Arg140, Gln143, Ala158, Glu161, Ser162, Ser165, Arg168, Leu268, Arg272, Ser275 and Gln279. Among them, Arg272, Glu161 and Arg168 have the highest frequency of occurrence and are key amino acid residues. The main binding sites of the hexapeptide SPPGER with TMC4 are Gln143, Glu161, Ser162, Arg168, Ser165, Arg272 and Gln279, among which Ser165 and Gln279 are key amino acid residues. By analyzing Figure 5 We found that the binding process is mainly through hydrogen bonding and electrostatic interactions. Among them, the hydrogen bonding ratio of the main active sites (Arg272, Glu161, Gln279, Arg168, Ser165) is 60% to 80%. Therefore, it can be assumed that hydrogen bonding is the most important non-covalent interaction between the peptide and the TMC4 receptor. The two-dimensional and three-dimensional best docking postures of the four peptides with TMC4 are shown in Figure 2. Figure 6 As shown. It can be observed that these four peptides all form hydrogen bonds with Ser165, Arg272 and Gln279, and form electrostatic interactions (salt bridges) with Glu161 and Arg168. The P residue benzene ring of the ligand SPDIY produces hydrophobic interactions with Arg272 through the alkyl group. In summary, we can prove that the four peptides SPDIY, DPSPR, QPGDY and SPPGER have significant salt-enhancing effects, which can lay the foundation for screening new salty taste enhancing peptides from peanut protein.

[0069] Determination of antioxidant activity

[0070] We evaluated the antioxidant activity of four salty taste enhancing peptides by studying their free radical scavenging abilities against DPPH and ABTS. The DPPH free radical scavenging rate and ABTS free radical scavenging rate were determined according to GB / T3900-2020 "DPPH and ABTS Method for Determination of Antioxidant Activity of Peptides". Figure 7As shown, QPGDY and DPSPR showed good DPPH free radical scavenging ability, among which the scavenging effect of QPGDY on DPPH was dose-related (p<0.05). At a concentration of 500μg / mL, the free radical scavenging rate of QY5 reached 60.33%. ABTS free radicals are also commonly used to evaluate antioxidant capacity. Both SPDIY and QPGDY showed ABTS free radical scavenging activity in the concentration range of 100 to 500μg / mL, and were significantly dose-dependent (p<0.05). At a concentration of 500μg / mL, the ABTS free radical scavenging rates of SPDIY (50.00%) and QPGDY (51.67%) were significantly higher than those of the other two peptides ( Figure 8 ). Therefore, it can be shown that the peptide QPGDY can scavenge at least one of the DPPH free radical and the ABTS free radical and has good antioxidant activity.

[0071] The specific embodiments described above further illustrate the objectives, technical solutions and beneficial effects of the present invention in detail. It should be understood that the above description is only a specific embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A peanut-derived salty taste enhancing peptide, characterized in that: The salty taste enhancing peptide is screened from peanut protein and binds to the TMC4 receptor protein. The salty taste enhancing peptide is composed of one of the following peptides: 1) the amino acid sequence is SPDIY, and the molecular weight is 593.63Da; 2) the amino acid sequence is DPSPR, and the molecular weight is 570.6Da; 3) the amino acid sequence is QPGDY, and the molecular weight is 578.57Da; 4) the amino acid sequence is SPPGER, and the molecular weight is 641.67Da.

2. The three salty taste enhancing pentapeptides according to claim 1, characterized in that: The peptide segment with the amino acid sequence of SPDIY, QPGDY and DPSPR was molecularly docked with the receptor TMC4, and the main binding sites were Lys136, Arg140, Gln143, Ala158, Glu161, Ser162, Ser165, Arg168, Leu268, Arg272, Ser275 and Gln279. Among them, Arg272, Glu161 and Arg168 appeared most frequently and were key amino acid residues.

3. The salty taste enhancing hexapeptide according to claim 1, characterized in that: The peptide segment with the amino acid sequence of SPPGER was molecularly docked with the receptor TMC4, and the main binding sites were Gln143, Glu161, Ser162, Arg168, Ser165, Arg272 and Gln279. Among them, Ser165 and Gln279 appeared most frequently and were key amino acid residues.

4. A salty taste enhancing peptide according to claim 1, characterized in that: The saltiness value of the peptide segment with the amino acid sequence of SPDIY is 4.04, the saltiness enhancement threshold is 0.64 mmol / L, and it can replace 6.32% NaCl; And / or, the peptide segment with the amino acid sequence of DPSPR has a saltiness value of 7.1, a saltiness enhancement threshold of 0.16 mmol / L, and can replace 87.1% of NaCl; And / or, the peptide segment with the amino acid sequence of QPGDY has a saltiness value of 4.42, a saltiness enhancement threshold of 0.33 mmol / L, and can replace 16.3% NaCl; And / or, the peptide segment with the amino acid sequence of SPPGER has a saltiness value of 4.49 and a saltiness enhancement threshold of 0.30 mmol / L, and can replace 18.2% NaCl.

5. A salty taste enhancing peptide according to claim 1, characterized in that: The peptide segment with the amino acid sequence of QPGDY can remove at least one of DPPH free radicals and ABTS free radicals and has antioxidant activity.

6. A method for screening salty taste enhancing peptides, characterized in that: Here are the steps: (1) Construct a TMC4 salty taste receptor protein model using the AlphFold2 online server and evaluate its reliability; The reliability evaluation: pLDDT score>70; the proportion of amino acid residues falling in the allowed region and the maximum allowed region in the whole protein is required to be higher than 90%, and the conformation of the model is considered reasonable and reliable; according to the Ramachandran diagram, the proportion of amino acid residues falling in the allowed region and the maximum allowed region in the whole protein is required to be higher than 90%; (2) Computer simulation of enzymatic hydrolysis of peanut protein to obtain enzymatic peptide fragments; (3) Predict the biological activity of enzymatic peptides with a sequence length of 4-8, screen out peptides with potential biological activity, predict their physicochemical properties, and screen out non-toxic and water-soluble peptides; The screening criteria for the biological activity prediction are: prediction score > 0.5; The screening criteria for toxicity prediction are: NonToxin; The screening criteria for water solubility prediction are: Good water solubility; (4) molecular docking the TMC4 receptor protein model with the peptide segment of step (3) to form a complex of the receptor protein and the small molecule ligand; The TMC4 receptor protein needs to remove water molecules and add hydrogen atoms under the CHARMM force field environment; The active position coordinates of the molecular docking are: X=-15.25, Y=11, Z=7.25, and the radius is 30; The molecular docking selects the largest cavity in the TMC4 receptor protein as the docking pocket; (5) Screening peptides with potential salt-increasing effects based on the molecular docking results and synthesizing the peptides by solid-phase synthesis; The screening: selecting peptides with low energy values ​​in CDOCKER energy; (6) Performing sensory evaluation and electronic tongue analysis on the synthesized peptide segment to obtain the salty taste enhancing peptide according to any one of claims 1 to 3.

7. Use of the salty taste enhancing peptide according to any one of claims 1 to 4 in the preparation of condiments and foods.

8. A product containing the salty taste enhancing peptide selected from peanut protein according to any one of claims 1 to 4.

9. Use of the salty taste enhancing peptide with antioxidant properties according to claim 5 in the preparation of condiments and foods.

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