Genomic Insights into the Role of cAMP in Carotenoid Biosynthesis: Enhancing β-Carotene Production in Escherichia coli via cyaA Deletion

The gamma-ray-induced random mutagenesis of an engineered β-carotene-producing <i>Escherichia coli</i> XL1-Blue resulted in the variant Ajou 45, which exhibits significantly enhanced β-carotene production. The whole-genome sequencing of Ajou 45 identified 55 mutations, notably including a reduction in the copy number of <i>cyaA</i>, encoding adenylate cyclase, a key enzyme regulating intracellular cyclic AMP (cAMP) levels. While the parental XL1-Blue strain harbors two copies of <i>cyaA</i>, Ajou 45 retains only one, potentially leading to reduced intracellular cAMP concentrations. This reduction may alleviate catabolite repression and redirect metabolic flux toward the β-carotene biosynthesis pathway. To validate this mechanistic insight, a targeted <i>cyaA</i> knockout was engineered in XL1-Blue, and its β-carotene production and growth phenotypes were compared with those of Ajou 45 and XL1-Blue. The findings demonstrated that a reduced <i>cyaA</i> copy number substantially enhances β-carotene biosynthesis by modulating cAMP-mediated regulatory networks. This study highlights the efficacy of integrating random mutagenesis with integrative genomic analysis for microbial strain engineering and presents a novel strategy for enhancing carotenoid production in <i>E. coli</i>.