Enhanced l-serine synthesis in Corynebacterium glutamicum by exporter engineering and Bayesian optimization of the medium composition

l-serine is a versatile, high value-added amino acid, widely used in food, medicine and cosmetics. However, the low titer of l-serine has limited its industrial production. In this study, a cell factory without plasmid for efficient production of l-serine was constructed based on transport engineering. Firstly, the effects of l-serine exporter SerE overexpression and deletion on the cell growth and l-serine titer were investigated in <i>Corynebacterium glutamicum</i> (<i>C. glutamicum</i>) A36, overexpression of <i>s</i> <i>erE</i> using a plasmid led to a 15.1% increase in l-serine titer but also caused a 15.1% decrease in cell growth. Subsequently, to increase the export capacity of SerE, we conducted semi-rational design and bioinformatics analysis, combined with alanine mutation and site-specific saturation mutation. The mutant E277K was obtained and exhibited a 53.2% higher export capacity compared to wild-type SerE, resulting in l-serine titer increased by 39.6%. Structural analysis and molecular dynamics simulations were performed to elucidate the mechanism. The results showed that the mutation shortened the hydrogen bond distance between the exporter and l-serine, enhanced complex stability, and reduced the binding energy. Finally, Bayesian optimization was employed to further improve l-serine titer of the mutant strain C-E277K. Under the optimized conditions, 47.77 g/L l-serine was achieved in a 5-L bioreactor, representing the highest reported titer for <i>C. glutamicum</i> to date. This study provides a basis for the transformation of l-serine export pathway and offers a new strategy for increasing l-serine titer.