# Immunomodulatory Peptides Derived from Tylorrhynchus heterochaetus: Identification, In Vitro Activity, and Molecular Docking Analyses

## TL;DR

This study identifies immunomodulatory peptides from Tylorrhynchus heterochaetus, showing they can activate immune cells and may be used in functional foods.

## Contribution

The study identifies and validates novel immunomodulatory peptides from Tylorrhynchus heterochaetus with potential for functional food applications.

## Key findings

- The <3000 Da fraction of hydrolyzed Tylorrhynchus heterochaetus showed strong immune-stimulating activity in macrophages.
- Three novel peptides (LPWDPL, DDFVFLR, LPVGPLFN) exhibited strong binding to TLR4/MD-2 receptors via molecular docking.
- Synthetic verification confirmed the peptides' safety and effectiveness in activating macrophages and releasing immune mediators.

## Abstract

Tylorrhynchus heterochaetus is an aquatic food with both edible and medicinal value in China. With a protein-rich body wall, it has strong potential for producing bioactive peptides. To explore its potential as a source of immunomodulatory peptides, in this study, flavor enzymes were selected as the optimal hydrolases, and the hydrolyzed products were subjected to ultrafiltration fractionation. The <3000 Da portion exhibited the most effective immune-stimulating activity in RAW 264.7 macrophages, enhancing phagocytosis and promoting the secretion of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and nitric oxide (NO) in a concentration dependent manner. Peptide omics analysis, combined with the activity and safety screened by bioinformatics, identified 43 candidate peptides. Molecular docking predicts that three novel peptides, LPWDPL, DDFVFLR and LPVGPLFN, exhibit strong binding affinity with toll-like receptor 4/myeloid differentiation factor-2 (TLR4/MD-2) receptors through hydrogen bonding and hydrophobic/π stacking interactions. Synthetic verification confirmed that these peptides were not only non-toxic to cells at concentrations ranging from 62.5 to 1000 µg/mL, but also effective in activating macrophages and stimulating the release of immune mediators. This study successfully identified the specific immunomodulatory peptides of the Tylorrhynchus heterochaetus, supporting its high-value utilization as a natural source of raw materials for immunomodulatory functional foods.