AcmA-mediated non-GMO probiotic engineering for functional jujube juice fermentation.

Probiotics are indispensable functional microorganisms in food fermentation with increasingly promising application prospects. Although genetically modified probiotics show great potential in improving the quality of fermented foods, their practical application is hampered by ethical controversies and safety concerns. To address these issues, the sacB gene encoding levansucrase was cloned from Bacillus atrophaeus in this study. Three non-genetically modified (non-GMO) engineered strains were constructed via the binding of AcmA to peptidoglycan through key hydrogen-bonding residues. In jujube juice fermentation, these strains catalyze the synthesis of levan with β - (2,6) glycosidic linkages. Untargeted metabolomics analysis demonstrated that this strategy rewires the sucrose metabolic pathway and activates secondary metabolic pathways, thereby promoting prebiotic synthesis and enabling systematic regulation of the microbial metabolic network. This innovative non-GMO technology circumvents potential risks associated with genetic modification and provides new avenues for the development of functional foods and the industrial application of probiotics.