Ribosome-Engineered Lacticaseibacillus rhamnosus Strain GG Exhibits Cell Surface Glyceraldehyde-3-Phosphate Dehydrogenase Accumulation and Enhanced Adhesion to Human Colonic Mucin

Differences in individual host responses have emerged as an issue regarding the health benefits of probiotics. Here, we applied ribosome engineering (RE) technology, developed in an actinomycete study, to Lacticaseibacillus rhamnosus GG (LGG). RE can effectively enhance microbial potential by using antibiotics to induce spontaneous mutations in the ribosome and/or RNA polymerase. In this study, we identified eight types of streptomycin resistance mutations in the LGG rpsL gene, which encodes ribosomal protein S12. Notably, LGG harboring the K56N mutant (LGG-MT K56N ) expressed high levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) on the cell surface compared with the LGG wild type (LGG-WT). GAPDH plays a key role in colonic mucin adhesion. Indeed, LGG-MT K56N significantly increased type A human colonic mucin adhesion compared to LGG-WT in experiments using the Biacore system. The ability to adhere to the colon is an important property of probiotics; thus, these results suggest that RE is an effective breeding strategy for probiotic lactic acid bacteria. IMPORTANCE We sought to apply ribosome engineering (RE) to probiotic lactic acid bacteria and to verify RE's impact. Here, we showed that one mutant of RE Lacticaseibacillus rhamnosus GG (LGG-MT K56N ) bore a GAPDH on the cell surface; the GAPDH was exported via an ABC transporter. Compared to the wild-type parent, LGG-MT K56N adhered more strongly to human colonic mucin and exhibited a distinct cell size and shape. These findings demonstrate that RE in LGG-MT K56N yielded dramatic changes in protein synthesis, protein transport, and cell morphology and affected adherence to human colonic mucin.