The acidogenicity of Streptococcus mutans is important for caries development. The antimicrobial peptide GH12 can affect the integrity of cellular membranes and the virulence factors of S. mutans. Combining GH12 and NaF (GF) efficiently controlled the development of caries, but its mechanisms remained unrevealed. This research intended to verify the effects of GF on the acidogenicity of S. mutans biofilms and to reveal the mechanisms. Lactic acid production assays and pH monitoring assays were conducted to investigate the regulatory effects of the GF treatment on the acidogenicity of S. mutans biofilms. RNA sequencing and bioinformatics analyses were conducted to screen the transcriptional profile affected by the GF treatment. The results demonstrated the GF group had significantly less lactic acid and maintained the broth's pH values above 5.0 for longer times. Thereafter, GO/KEGG enrichment analyses and RT-qPCR validation revealed that the GF treatment mainly restrained the expression of genes related to the carbohydrates' internalization and metabolism. Compared with NaF, the GF treatment further downregulated the carbohydrates transportation genes. Moreover, compared with GH12, the GF treatment affected the membrane's integrity more significantly. Generally, GF treatment could arrest the acidogenicity of S. mutans biofilms, mainly through suppressing carbohydrates transportation and inhibiting overall metabolism.