Our previous study indicated that ethanol-induced intracellular extracts (E-IEs) of Lactococcus lactis subsp. Lactis IL1403 (L. lactis IL1403) alleviated hangovers more effectively in mice than untreated intracellular extracts (U-IEs), but the material basis was unclear. Considering that stress-related proteins might play a significant role, the effects of ethanol induction on probiotic properties of L. lactis IL1403 and the associated stress response mechanism were initially explored in this study. E-IEs of L. lactis IL1403 showed better biological activities, significantly increased bacteria survival rates in oxidative stress environments, increased ADH activity, and enhanced proliferation in RAW264.7 and AML-12 cells. Proteomic analyses revealed that 414 proteins were significantly changed in response to ethanol induction. The expression of proteins involved in the universal stress response, DNA repair, oxidative stress response, and ethanol metabolism was rapidly upregulated under ethanol stress, and quantitative real-time PCR (qRT-PCR) results were consistent with proteomic data. KEGG pathway analysis indicated that citrate metabolism, starch and sucrose metabolism, and pyruvate metabolism were significantly enriched during ethanol stress to increase energy requirements and survival rates of stressed cells. Based on this observation, the active induction is an effective strategy for increasing the biological activity of L. lactis IL1403. Exploring the molecular mechanism and material basis of their functions in vivo can help us understand the adaptive regulatory mechanism of microorganisms.