Taste-Active Peptides from Triple-Enzymatically Hydrolyzed Straw Mushroom Proteins Enhance Salty Taste: An Elucidation of Their Effect on the T1R1/T1R3 Taste Receptor via Molecular Docking

Abstract

The objective of our study was to analyze and identify enzymatic peptides from straw mushrooms that can enhance salty taste with the aim of developing saltiness enhancement peptides to reduce salt intake and promote dietary health. We isolated taste-related peptides from the straw mushroom extract using ultrafiltration and identified them using UPLC-Q-TOF-MS/MS. The study found that the ultrafiltration fraction (500-2000 Da) of straw mushroom peptides had a saltiness enhancement effect, as revealed via subsequent E-tongue and sensory analyses. The ultrafiltration fractions (500-2000 Da) were found to contain 220 peptides, which were identified through UPLC-Q-TOF-MS/MS analysis. The interaction of these peptides with the T1R1/T1R3 receptor was also assessed. The investigation highlighted the significant involvement of Asp223, Gln243, Leu232, Asp251, and Pro254 in binding peptides from triple-enzymatically hydrolyzed straw mushrooms to T1R1/T1R3. Based on the binding energy and active site analysis, three peptides were selected for synthesis: DFNALPFK (-9.2 kcal/mol), YNEDNGIVK (-8.8 kcal/mol), and VPGGQEIKDR (-8.9 kcal/mol). Importantly, 3.2 mmol of VPGGQEIKDR increased the saltiness level of a 0.05% NaCl solution to that of a 0.15% NaCl solution. Additionally, the addition of 0.8 mmol of YNEDNGIVK to a 0.05% NaCl solution resulted in the same level of saltiness as a 0.1% NaCl solution.

Keywords: T1R1/T1R3; UPLC-Q-TOF-MS/MS; molecular docking; peptides from triple-enzymatically hydrolyzed straw mushroom proteins; straw mushroom.

Figure 1

Results of solids content and the degree of hydrolysis (DH) after treatment with three enzymatic methods. The line matches the left y-axis (solids content), and the bar matches the right y-axis (DH). According to a one-way ANOVA, the same letter of the marker indicates that there is no statistical difference between the amounts of the solids content and DH in the supernatant obtained using different enzymatic methods (n = 3, p < 0.05).

Figure 2

(A,C) are the results of the E-tongue evaluation, and (B,D) are the results of the sensory evaluation. The substrate in (A,B) is the enzyme supernatant, and the substrate in (C,D) is a mixture of a 0.5% NaCl solution and the enzyme supernatant. The dotted line in (B) indicates the score for the ability to taste saltiness. The dotted line in both (C,D) indicates the score for the 0.5% NaCl solution. According to a one-way ANOVA, the same letter of the marker indicates that there is no statistical difference between the amounts of the E-tongue value and sensory score in the supernatant obtained using different enzymatic methods. (n = 3, p < 0.05).

Figure 3

Results of the 2-AFC experiment. The vertical axis indicates the number of assessors who agreed that the different ultrafiltration fractions were saltier. The dashed line indicates the minimum number of agreement judgments required to identify the saltier samples in a comparison test at the α = 0.05 level. According to a one-way ANOVA, the marks with the same letter indicate that there is no statistical difference between the number of assessors at different concentrations of the salt solution (n = 3, p < 0.05).

Figure 4

The gLMS scores (A) and the change of saltiness intensity (B) under three ultrafiltration fractions. According to a one-way ANOVA, the same letter of the marker indicates that there is no statistical difference between the amounts of the gLMS scores in different ultrafiltration fractions (n = 3, p < 0.05).

Figure 5

Statistics of the active sites of eight taste peptides interacting with the T1R1/T1R3 umami receptor.

Figure 6

Two-dimensional diagram of the docking of DFNALPFK (A), DKLHEGIK (B), GVGPFDDDR (C), IDNEPEFRWA (D), IGDEAAENRV (E), SEHEENGYAV (F), VPGGQEIKDR (G), and YNEDNGIVK (H) with T1R1/T1R3.

Figure 7

The saltiness enhancement characteristics of synthetic peptides. The bar matches the left y-axis (E-tongue), and the line matches the right y-axis (sensory evaluation). According to a one-way ANOVA, the same letter of the marker indicates that there is no statistical difference between the amounts of the E-tongue and sensory evaluation in different synthetic peptides (n = 3, p < 0.05).

Figure 8

Dose–feedback results for three synthetic peptides. The green line represents VP10, the blue line represents YN9, and the orange line represents DF8. Using a one-way ANOVA (n = 3, p < 0.05).