Transcriptome-Guided Analysis and Application of the Mechanisms Influencing PamyE Promoter Activity

The commonly used P_amyE promoter in Bacillus amyloliquefaciens industrial enzyme production is highly susceptible to carbon catabolite repression (CCR). To address this limitation, a modified promoter, P_ΔCRE2, was engineered by deleting the carbon-responsive element (CRE), which mitigated CCR. However, its activity still varied across different carbon sources, indicating the presence of regulatory mechanisms beyond the canonical CcpA-CRE pathway, referred to as additional carbon catabolite repression (ACCR). Transcriptomic analysis identified CcpA, Spo0A, and DegU as key regulatory factors, and functional validation demonstrated that deletion of ccpA combined with overexpression of spo0A and degUS enhanced P_ΔCRE2 activity. The resulting engineered strain BA4 exhibited a 120% increase in aminopeptidase activity, highlighting the potential of promoter optimization combined with metabolic rewiring to improve carbon-source utilization.