Multimodular Pathway Engineering of Bacillus amyloliquefaciens for Intensifying Iturin A Production

Iturin A is a potent lipopeptide with broad application potential. However, its microbial production remains constrained by limited precursor availability and metabolic inefficiencies. In this work, a modular metabolic engineering strategy was employed in Bacillus amyloliquefaciens to systematically enhance the biosynthetic capacity of multiple precursor pathways. Fatty acid synthesis was improved by deleting the transcriptional repressor fapR, overexpressing the key gene fabG, and disrupting mmgA, which is involved in acetyl-CoA metabolism. To increase the availability of proline and serine, their catabolic genes ( fadM and sdaAB ) were deleted, while the corresponding biosynthetic genes ( proBA and serC ) were overexpressed. Furthermore, the branched-chain amino acid pathway was reinforced by overexpressing the key gene ilvD and deleting the aminotransferase gene gabT . Nitrogen acquisition was also further enhanced by overexpressing the opp operon encoding an oligopeptide transporter. The resulting engineered strain, ITUz21, achieved an iturin A titer of 7.62 g/L and a productivity of 0.14 g/L/h in a 7.5 L bioreactor, representing the highest level reported to date. These findings demonstrate the effectiveness of coordinated precursor pathway engineering for improving microbial iturin A biosynthesis.