There are multiple lines of evidence that lactic acid bacteria (LAB) exert cancer-preventive effects. However, the underlying mechanisms are poorly understood. In the present study we found that the cytoplasmic fraction of Lactococcus lactis ssp. lactis American Type Culture Collection (ATCC) 7962 exerted the strongest antiproliferative effects (half-maximal inhibitory concentration (IC50) = 17 μg/ml) in SNU-1 human stomach cancer cells and arginine deiminase (ADI; EC 3.5.3.6) activity. We also cloned, expressed and purified ADI from L. lactis ssp. lactis ATCC 7962 (LADI). Both purified ADI from L. lactis (PADI; IC50 = 2 μg/ml) and recombinant ADI originating from LADI (IC50 = 0·6 μg/ml) inhibited the proliferation of SNU-1 cells. LADI induced G0/G1-phase arrest, sub-G1 accumulation, DNA condensation and DNA fragmentation in SNU-1 cells. 4′,6-Diamidino-2-phenylindole (DAPI) staining and DNA fragmentation data provide evidence that LADI induces apoptosis in SNU-1 cells. LADI increased the expressions of p53 and p27Kip1, and decreased the expressions of cyclin D1, c-myc and Bcl-xL in SNU-1 cells. However, LADI had no effects on the expressions of p21Cip1 and Bcl-2. Collectively, these data indicate that ADI induces apoptosis and G0/G1-phase arrest of SNU-1 cells, which might contribute to the chemopreventive potential of LAB.