关键词: 咸味肽；TMC4受体；AlphaFold3；分子对接；亲水性氨基酸

Abstract: This study employed the SeqLogo method to examine the sequence characteristics of salty peptides. Utilizing the AlphaFold3 de novo folding approach, a model of the salty receptor transmembrane channel like 4 (TMC4) was constructed. Concurrently, molecular simulation and frontier molecular orbital (FMO) calculation were applied to elucidate the interaction mechanism between salty peptides and the TMC4 receptor. Results indicated that under physiological pH conditions, salty peptides were rich in charged amino acids, with hydrophilic residues predominantly occurring at the N- or C-terminus of most sequences. Molecular docking analysis revealed that long-chain peptides (8−11 amino acids) exhibited significantly higher affinity for the TMC4 receptor than short-chain peptides. Furthermore, the saltiness intensity of salty peptides was negatively correlated with the docking score of the TMC4 receptor (P < 0.01). Salty peptides primarily bound to the TMC4 receptor via hydrogen bonds, thereby exerting their salty effect. ALA401, PHE405, LYS412, ARG437, VAL495, GLN524, GLN527, and GLU531 constituted key binding sites on the TMC4 receptor. Meanwhile, hydrophilic amino acids in salty peptides and TMC4 played pivotal roles in their interaction. Electrostatic potential energy and loss-of-function mutation analyses confirmed Arg as a critical amino acid within salty peptides. Molecular dynamics simulations further elucidated the binding stability, interaction forces, and key binding sites between salty peptides and TMC4. FMO calculations identified amino acid residues such as Asp and Glu as primary active sites for salty peptide binding to TMC4, with small energy gaps tending to correspond to low docking scores. Salty peptides with lower binding energies to the TMC4 receptor tended to demonstrate stronger perceived saltiness intensity in sensory evaluation. These findings facilitate the identification of potential salty peptides, provide a theoretical foundation for establishing computationally-driven high-throughput screening systems, and offer innovative strategies for low-salt food development.

Key words: salty peptides; TMC4 receptor; AlphaFold3; molecular docking; hydrophilic amino acids