Development of a High Anthraquinone-Producing <i>Escherichia coli</i> Strain Using Malonyl-CoA Supply Pathway Engineering

Anthraquinones are valuable compounds that are traditionally used as natural pigments and have diverse pharmacological activities, including antimicrobial and anticancer effects. In this study, we aimed to enhance the production of 1,3,5-trihydroxyanthraquinone (AQ256) using <i>Escherichia coli</i> (<i>E. coli</i>) as a host. AQ256 is biosynthesized from eight malonyl-CoA molecules via the type II polyketide synthase pathway. However, previous studies have reported very low production levels of AQ256 in <i>E. coli</i> (approximately 2.5 mg/L), mainly because of limited malonyl-CoA availability. To address this, we introduced a heterologous malonate assimilation pathway and reinforced the endogenous malonyl-CoA biosynthesis pathway. An <i>E. coli</i> strain harboring AQ256 biosynthetic genes from <i>Photorhabdus laumondii</i> TTO1 produced only 1.3 mg/L AQ256. Upon introducing the malonate assimilation pathway and cultivating in malonate-supplemented Luria-Bertani medium, production increased to 3.8 mg/L. Further enhancement of the endogenous malonyl-CoA supply through the coexpression of pantothenate kinase and acetyl-CoA carboxylase resulted in strain AQ-04, which produced 12.3 mg/L AQ256. Optimization of cultivation conditions enabled AQ-04 to achieve 23.9 mg/L AQ256, a 9.6-fold increase compared to previous studies. Our results demonstrate that the combination of introducing a malonate assimilation pathway and enhancing native malonyl-CoA supply is a highly effective strategy for increasing malonyl-CoA availability. This approach is promising for the biosynthesis of a wide range of malonyl-CoA-derived compounds.