The oral cavity is considered an extra-gastric reservoir for Helicobacter pylori (H. pylori) and oral H. pylori can contribute to the gastric eradication inability and recurrence. However, the oral environment is not ideal for H. pylori survival, and the factors promoting oral colonization and survival of H. pylori have not been elucidated. In this study, we explored the effects of extracellular polysaccharides (EPS), the fundamental building blocks of dental Streptococcus mutans (S. mutans) biofilm, on H. pylori colonization and drug resistance in the oral cavity, as well as stomach. In the co-culture system of H. pylori Sydney strain (SS1) and three S. mutans biofilms with different EPS contents (UA159 wild-type, UA159ΔgtfB, UA159ΔgtfBC), it was found that the adhesive force between SS1 and biofilms increased correspondingly with the increase in EPS content. Moreover, with the increase in EPS content of biofilms, the number of colonized SS1 increased. Proteome analysis revealed that SS1 co-cultured with UA159 biofilm exhibited 149 differentially expressed proteins compared to that co-cultured with UA159ΔgtfB biofilm, with significant enrichment in β-lactamase activity pathway. SS1 co-cultured with UA159 biofilm exhibited 154 differentially expressed proteins compared to that co-cultured with UA159ΔgtfBC biofilm, with significant enrichment in β-lactamase activity, aminoglycoside nucleotidyltransferase activity and antioxidant activity pathways. Both in vivo and in vitro, EPS synthesized by glucosyltransferases (Gtfs) surrounding SS1 was verified to protect SS1 against β-lactam and aminoglycoside antibiotics. These findings demonstrated that S. mutans biofilms mediate oral adhesion, colonization, and antibiotic resistance of H. pylori through a Gtfs-driven EPS biosynthesis mechanism.