Elucidating metabolic mechanisms underlying the influence of specific growth rate on alkaline protease synthesis in Bacillus licheniformis through combined omics and computational modeling analysis.

Specific growth rate is a crucial physiological parameter in fermentation. This study integrates transcriptomics, metabolomics and genome-scale metabolic modeling to uncover the physiological changes in Bacillus licheniformis induced by specific growth rate during fed-batch fermentation. The results showed that increasing growth rates significantly enhanced both cell growth and alkaline protease activity. Transcriptomic analysis revealed significant upregulation of genes related to energy supply, phosphate response and protease synthesis regulation at elevated growth rates. Metabolic flux analysis demonstrated that the alkaline protease accumulation was promoted by upregulating central carbon metabolism and increasing the synthesis of amino acids in high demand, including alanine, glycine, valine, and lysine. Scaling to a 50 L bioreactor further optimized alkaline protease activity to 126,786.54 U/mL at a growth rate of 0.04 h. This study elucidates metabolic and regulatory drivers of growth-coupled protease production, providing insights into its optimization for industrial scale fermentation.