With the growing antibiotic resistance of Staphylococcus aureus, it is urgent to discover new antimicrobials to solve this problem. By bacterial growth based high-throughput screening, we identified FDA-approved drug T-2307 with effective antimicrobial activity against S. aureus. This study aims to explore the model of action of the compound by proteomics analysis and other microbiological methods. The data would provide us information about the regulated proteins, and hence help us further study the underlying antimicrobial mechanisms by the compound.