Structural characterization and immunomodulatory activity of a glycogen from Meretrix lyrata

Marine shellfish are not only important food ingredients in human diets, but also their tissues contain polysaccharides with significant bioactivities and pharmacological effects. In this study, a glucan (MLP60-1) was successfully isolated from the soft tissue of Meretrix lyrata. Structural characterization through monosaccharide compositions, methylation analysis, acid and enzymatic hydrolysis, mass spectrometry, and nuclear magnetic resonance spectroscopy suggested that MLP60-1 is a highly branched glycogen defined by a branching frequency of approximately one branch per 3.3 glucose residues. The backbone of MLP60-1 consists of →4)-α-D-Glcp-(1→ residue, with branches substituted at the C-6 position comprising →4,6)-α-D-Glcp-(1→, α-D-Glcp-(1→, →6)-α-D-Glcp-(1→, and →4)-α-D-Glcp-(1→ residues. Subsequently, in vitro immunomodulatory assays indicated that MLP60-1 significantly enhanced nitric oxide (12.5 μg/mL, p < 0.05) and reactive oxygen species production and stimulated the secretion of tumor necrosis factor-α, interleukin-6, and interleukin-1β in RAW264.7 cells. The molecular mechanism study was demonstrated that MLP60-1 promotes macrophage activation via the TLR4/MyD88/NF-κB signaling pathway. Subcellular localization of fluorescently labeled derivatives, siRNA interference, microscale thermophoresis assay (K_d = 9.89 ± 1.50 μmol/L), and molecular docking analysis further suggested TLR4 is likely to plays a potential crucial role as membrane protein target mediating MLP60-1-induced macrophage activation and immunoregulation. Collectively, these findings suggest that MLP60-1 exhibits potential immunomodulatory properties, which may support its future exploration as a natural functional food ingredient.