L-Lysine production from glucose and chitin monomers using engineered Vibrio natriegens

Abstract Despite its industrial importance, microbial L-lysine production has largely been confined to classical producer strains, leaving the fast-growing, non-pathogenic marine bacterium V. natriegens mostly unexplored as an emerging biosynthetic platform. In this work, we established an L-lysine-overproducing V. natriegens DSM759 strain through a stepwise rational engineering strategy targeting the native biosynthetic pathway. Building on our prior systems-level analysis of the strain’s genetic and regulatory architecture, we identified key metabolic bottlenecks and targeted interventions to relieve them. The central intervention was relief of feedback inhibition on the two key regulatory enzymes, aspartate kinase (AK, lysC ) and dihydrodipicolinate synthase (DHDPS, dapA ). Transferring feedback resistance substitutions previously characterized in E. coli to conserved positions in the V. natriegens DSM759 enzymes produced L-lysine-insensitive variants with kinetic parameters comparable to the corresponding wild-type enzymes. Among the tested configurations, the strain co-expressing Vn.lysC2 and Vn.dapA1 :E84T reached the highest L-lysine titer (9.0 ± 0.6 mM) and yield (0.11 ± 0.01 mol Lys mol Glc −1 ), whereas overexpression of additional L-lysine pathway genes provided no further benefit. Exploiting the host’s substrate range, we further demonstrated L-lysine synthesis from the chitin-derived amino sugar N-acetylglucosamine (0.09 ± 0.00 mol Lys mol GlcNAc −1 ), highlighting the potential to valorize chitin-rich waste streams from the seafood industry. This work establishes a minimal rational strategy for L-lysine biosynthesis in V. natriegens DSM759 and positions it as a candidate platform for sustainable amino acid production. Key points • E. coli-derived rational engineering strategies were transferable to V. natriegens • Native pathway deregulation was sufficient to enable L-lysine production • Chitin-derived N-acetylglucosamine supports L-lysine synthesis