Novel immunomodulatory peptides identified from Meretrix meretrix L. hydrolysates through in vitro cell experiments, in silico analysis, molecular docking, and dynamic simulation

Our study aimed to identify novel immunomodulatory peptides derived from the hydrolysates of Meretrix meretrix L. (MMLHs). Following an immunomodulatory activity- guided isolation using size-exclusion chromatography, 58 peptides with molecular weights ranging from 529.65 Da to 1873.39 Da were identified. Comprehensive in silico analysis, incorporating multiple predictive tools, led to the selection of two peptides, SLLELDL (SL) and LEKNKDPLNET (LEK) for synthesis, followed by subsequent activity validation. Both SL and LEK significantly activated RAW264.7 macrophages, enhancing cell proliferation, promoting phagocytosis, and increasing the secretion of nitric oxide and cytokines, including TNF-α and IL-1β. Molecular docking studies revealed strong binding affinities of SL and LEK to the TLR4-MD2 receptor, primarily through hydrophobic interactions, with binding energies of −6.1 kcal/mol and −6.0 kcal/mol, respectively. The root mean square deviations (RMSD) and radius of gyration (Rg) values indicated that compared to the LEK-TLR4-MD2 complexes, SL-TLR4-MD2 complexes had more stable and compact conformations. Overall, this study is the first to demonstrate that Meretrix meretrix L. is a promising source of immunomodulatory peptides, capable of regulating inflammatory cytokine secretion and enhancing immune system function. • 58 peptides with MW ranged from 529.65 Da to 1873.39 Da were identified from MMLHs. • Two peptides, SLLELDL (SL) and LEKNKDPLNET (LEK), were selected after screening and verifying. • SL and LEK enhanced proliferation, phagocytosis, and cytokine secretion of macrophages. • SL and LEK bound to TLR4-MD2 receptor with high affinities via hydrophobic interactions. • Molecular dynamics simulations show that SL-TLR4-MD2 complex is more stable and compact than LEK.