Enhancing surfactin production in Bacillus subtilis: Insights from proteomic analysis of nitrate-induced overproduction and strategies for combinatorial metabolic engineering

Surfactin biosynthesis in Bacillus subtilis is intricately regulated by environmental conditions. In the present study, addition of nitrate, a nitrogen source, increased the production of surfactin in B. subtilis ATCC 21332, whereas its absence resulted in minimal or no surfactin production. Proteomics revealed the mechanism underlying nitrate-induced surfactin overproduction, identifying three key differential proteins (preprotein translocase subunit SecA, signal recognition particle receptor FtsY, and cell division adenosine triphosphate-binding protein FtsE) relevant to surfactin transport and regulation. Combinatorial metabolic engineering strategies (enhanced nitrate reduction, fatty acid hydroxylation, rational transporter engineering, and feeding) led to a 41.4-fold increase in surfactin production compared with the initial production in the wild-type strain. This study provides insights into the molecular mechanism of nitrate-induced surfactin overproduction and strategies to enhance the performance of surfactin-producing strains.