CyLip-20 is a cyclic lipid antimicrobial peptide targeting both Gram-positive and Gram-negative bacteria. It exhibits low hemolytic activity and mild in vivo toxicity. CyLip-20 disrupts the integrity of bacterial outer, inner, and cytoplasmic membranes by binding to bacterial lipopolysaccharides (LPS), leading to membrane permeabilization, depolarization, and leakage of cellular contents, while also inhibiting bacterial biofilm formation. In animal models, CyLip-20 reduces bacterial load in MRSA-infected mouse skin wounds, promotes wound healing, decreases the levels of inflammatory factors, and reduces inflammatory cell infiltration. CyLip-20 is applicable in research related to MRSA skin wound infections.