Antibiotic-resistant Staphylococcus aureus is beginning to pose a social issue. Thus, there is an urgent need for the development of effective anti-staphylococcal agents to eradicate antibiotic-resistant S. aureus in food systems and to treat the pathogen in clinical areas. To address this need, lactic acid bacteria (LAB) from kimchi were screened for the production of anti-staphylococcal bacteriocin. From this screening, a bacteriocin generated by the MK3 strain, which was identified by 16S rRNA gene sequence analysis as Enterococcus faecium, demonstrated antimicrobial activity against an S. aureus strain, and was designated enterocin MK3. Enterocin MK3 also demonstrated activity against other gram-positive bacteria, including several LAB and Listeria monocytogenes, but not gram-negative Escherichia coli. The molecular mass of enterocin MK3 was estimated as approximately 6.5 kDa on an SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) gel.