Dynamic Pathway Regulation Using a Quorum-Sensing Circuit to Improve the Production of L-Homoserine in <i>Escherichia coli</i>

L-Homoserine has diverse applications in the fields of agrochemicals, pharmaceuticals, and animal feed; therefore, microbial fermentation using engineered cell factories has attracted widespread and intense attention. In this study, the nonauxotrophic strain for higher L-homoserine production was developed based on the previously constructed strain <i>Escherichia coli</i> HS. First, key genes involved in the biosynthesis pathways of essential amino acids were replenished with different strategies to address the growth deficiencies. Subsequently, carbon flux through the l-aspartate to L-homoserine was amplified by <i>thrA</i> overexpression. Furthermore, the supply of NADPH and ATP was optimized to synergistically enhance L-homoserine biosynthesis. Finally, the quorum sensing (QS) system <i>esaI</i>/<i>esaR</i> from <i>Pantoea stewartii</i> was introduced to dynamically regulate the carbon flux of l-threonine biosynthesis. And the results indicated that optimizing the regulatory efficiency of the QS system triggered autonomous downregulation of <i>thrB</i> during the high-cell-density phase, achieving a balanced metabolic flux competition between the l-threonine and L-homoserine biosynthesis pathways. The QS-regulated strain <i>E. coli</i> HS27/PA-P7QS produced 101.81 g/L L-homoserine with a yield of 0.41 g/g glucose after 96 h of fermentation in a 5-L bioreactor. This study demonstrates the feasibility of applying the QS system in <i>E. coli</i> for metabolic flux control, thereby providing novel insights into the rational design of amino acid biosynthesis pathways.