A strain, Rahnella aquatilis L103, which was isolated from the soil around the roots of mushrooms, produced antibacterial protein though fermentation. This protein has broad antibacterial spectrum towards gram-positive and gram-negative bacteria. The protein has a significant antioxidant capacity on scavenging ABTS(+center dot) (82.5%, 120 mu g mL(-1)), DPPH center dot (64.1%, 600 mu g mL(-1)) and OH center dot (60.1%, 750 mu g mL(-1)) as well. The protein was considered to be safe within concentration of 150 mu g mL(-1) as shown by MTT results and was applied to beef refrigerated preservation. The treatment group presented lower total viable counts (TVC), total volatile basic nitrogen (TVB-N), 2-thiobarbituric acid (TBA), meanwhile, higher sensory score and longer shelf life (2 days) compared with the control group (P < 0.05). The results indicated that the protein under safe concentration was beneficial to beef preservation.