T1R1-VFT-Based Sensor Construction And Evaluation Of Umami Substance Interactions

As one of the five basic taste sensations,umami is favored in Chinese food culture for its unique and wonderful"taste bud-pleasing"appeal,and is an important factor in evaluating food flavor.The perception of umami in mammals is induced by the interaction between umami substances and umami receptors,and is perceived by the brain through cascade signaling.Different kinds of umami substances could induce umami perception,when multiple umami substances are present,they act on the umami receptors at the same time,resulting the synergistic effects of umami substances and further improving the food flavor.Therefore,examining the synergistic effects of umami substances is important in the flavoring industry.Therefore,this study constructed a biosensor for evaluating umami taste synergism,as well as verified the key amino acid residues by molecular simulation.The main study contents and results are as follows:（1）The N-Venus Flytrap domain of the umami taste receptor protein（T1R1-VFT）was obtained by heterologous expression as the recognition progenitor,Gold nanoparticles（Au NPs）were coupled into Au NPs-T1R1-VFT as the carrier.Subsequently,the bimetallic nanocomposite（Mo S₂-Pt Pd）was prepared by a modified method without heating,and the bimetal bionic receptor sensor was constructed based on its large specific surface area and high catalytic efficiency as the substrate material of the sensor.The sensor is characterized by the change of electrical signal,which could effectively identify the binding of T1R1-VFT and umami molecules.（2）The constructed sensor had wide detection range(10^-13-10^-5M),low detection limit（0.03 p M）and excellent linearity（R²>0.99）with good specificity,stability and reproducibility in detection of different types of umami substances（amino acids,nucleotides,organic acids,umami peptides）.Compared with traditional electronic tongue,the sensor possessed multiple features of human taste system and could effectively identify the umami taste synergism.The difference of umami taste synergism at different concentrations and ratios was also explored,and the results showed that the synergistic effect was more obvious at lower concentrations and balanced addition of multiple umami taste substances（p<0.05）.The signal response was influenced by the level of receptors binding.Meanwhile,it could also accurately detect the synergistic effect in complex matrices,showing good specificity and interference resistance.（3）Based on predictive scoring model i Umami-SCM and machine learning discriminative umami/bitter dichotomous model Umami＿YYDS,the umami peptides were scored and the identified umami peptides were selected by combining molecular weight,sensory evaluation and other indicators.The molecular simulation technique was used to construct the model of umami peptides and T1R1-VFT for docking,the results showed that the affinity of the two was high(binding energy<-7 kcal mol^-1)and the structure of the complex was stable;when multiple umami peptides were docked to T1R1-VFT simultaneously,the affinity was further increased by the synergistic effect of the ligand(binding energy<-8 kcal mol^-1).The interaction process between umami peptides and T1R1-VFT were analyzed by biolayer interferometry technique,and it was found that the umami peptides could complete the association and dissociation process with T1R1-VFT in solution.When multiple umami peptides interacted with T1R1-VFT solution at the same time,the kinetic equilibrium constant decreased,the affinity increased and the synergistic effect was significant（p<0.05）.In summary,this work from simulation to practical validation,evaluating the effect of umami synergy from multiple perspectives,identifying the key sites of umami synergy and analyzing the kinetic process of umami synergy,providing theoretical support for the high value utilization of umami flavor base materials and the development of specialty condiments products.