PNMGL2 exerts antibacterial effects through dual mechanisms: membrane disruption and biofilm inhibition.
PNMGL2 employs an α-helical structure combined with positive charge characteristics to electrostatically bind to the negatively charged bacterial membrane surface, subsequently inserting into the phospholipid bilayer to form pores. Bactericidal kinetics revealed that PNMGL2 eradicates 99% of Listeria monocytogenes at its minimum inhibitory concentration (MIC). Membrane integrity assays demonstrated a significant increase in propidium iodide (PI) staining rates in treated groups, alongside lactate dehydrogenase (LDH) leakage reaching 0.01 mmol/L. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) observations confirmed membrane invagination, wrinkling, and pore formation. Increased membrane permeability led to substantial efflux of potassium ions, phosphate, ATP, nucleic acids, and proteins, directly disrupting energy metabolism and genetic material stability.

PNMGL2 exhibited concentration-dependent inhibition of Listeria monocytogenes biofilm formation, reducing bacterial adhesion to food-processing surfaces and mitigating drug resistance. This mechanism may involve interference with quorum sensing systems or extracellular polymeric substance (EPS) synthesis, though exact pathways require further validation.

Hemolysis assays showed no erythrocyte lysis even at 4× MIC concentrations, indicating low mammalian cell toxicity. This safety profile stems from PNMGL2's specific targeting of bacterial membranes, avoiding interactions with eukaryotic cell membranes—a critical advantage for food preservative applications.

As a naturally derived antimicrobial, PNMGL2 addresses concerns about chemical preservative resistance through its dual-action mechanism. Future work should employ omics technologies to identify molecular interaction targets with membrane proteins or lipids. Additionally, its antimicrobial stability in complex matrices (e.g., dairy and meat products) and impacts on sensory quality must be evaluated to advance industrial applications.