Abstract: The rapid emergence of multidrug-resistant bacterial pathogens, particularly methicillin-resistant Staphylococcus aureus (MRSA), underscores the critical discovery for new antibiotics with novel scaffolds. We identified lead compound LXW933, which exhibits promising anti-MRSA activity and features a unique 1,2,4-oxadiazole scaffold, from our phidianidine-based compound library. Based on this structure, a Function-Oriented Synthesis (FOS) strategy had been conducted to afford a series of phidianidine derivatives for their antibacterial activity evaluation. Compounds 24a exhibited excellent anti-MRSA efficacy with an MIC value of 1.5 μg/mL and low toxicity against HeLa cells. Further mechanistic study showed that 24a effectively inhibits biofilm formation and exerts its antibacterial effect by disrupting the bacterial cell membrane. This research provides valuable insights for the discovery of antibacterial drug leads derived from oxadiazole-containing marine alkaloids, paving the way for new strategies to combat antimicrobial resistance.