Biosynthesis of the Functional Component Spermidine from <i>Bacillus amyloliquefaciens</i> by Iterative Integration Expression

Spermidine finds broad applications across both the nutraceutical and biomedical sectors. In this study, key regulatory genes affecting spermidine synthesis and efficient integration sites were identified to construct a chassis strain for green and sustainable spermidine production. First, the expression of <i>argJ</i> was increased, and the protein SAM2 was mutated to promote the synthesis of spermidine. Second, positional effects were examined in <i>Bacillus amyloliquefaciens</i>. Concurrently, bioinformatics analysis was conducted to uncover transport proteins Blt, YvdR, and Mta, as well as other key genes <i>tcyJ</i>, <i>yxeM</i>, <i>appC</i>, <i>yngA</i>, and orf03307 that affect spermidine synthesis. Ultimately, strain PM13 was constructed through the iterative integration of key genes, achieving a spermidine titer of 396.92 mg/L, 10.34 times higher than strain PM1. Furthermore, xylose fed-batch fermentation increased spermidine titer to 1.69 g/L, setting a new shake flask production record. In conclusion, this study amassed genetic resources and developed an integrated strain for efficient, stable spermidine synthesis.