Dynamic control strategies in metabolic engineering enable on-demand gene expression in genetically modified microorganisms, reducing unnecessary gene expression, optimizing cellular resource allocation, and boosting target product yields. Quorum sensing and riboregulator-based genetic circuits are emerging tools in synthetic biology, offering autonomous and precise gene regulation in engineered microorganisms.
This research integrates the LuxI/LuxR quorum sensing system with a riboregulator-based genetic circuit to achieve completely autonomous regulation in metabolic pathways. Using our predictive model-based antisense RNA (asRNA) design [1], we downregulated the
E. coli pfk-1 gene and combined it with the small transcription activating RNAs (STARs) to construct a genetic inverter. Then, an autoinducer production module was added to the circuit to enable the engineered strain to autonomously rewire the metabolic flux from the glycolytic pathway to the target production pathway. This circuit design was first applied to optimize the heterologous Myo-inositol production pathway, improving yield significantly. Subsequently, the circuit was extended to regulate the downstream glucaric acid pathway [2], incorporating three orthogonal STAR variants to balance metabolic flux by varying the mRNA level and further optimizing the production. These results demonstrate that autoregulated riboregulator-based circuits can effectively enhance dynamic gene expression control and improve the overall productivity of complex metabolic pathways in engineered microorganisms.
[1] YJ Lee, SJ Kim, MB Amrofell and TS Moon. Establishing a multivariate model for predictable antisense RNA-mediated repression. ACS Synth. Biol. 8, 45–56 (2019)
[2] TS Moon, S-H Yoon, A Lanza, J Roy-Mayhew and KJ Prather. Production of Glucaric Acid from a Synthetic Pathway in Recombinant Escherichia coli. Appl. Environ. Microbiol. 75, 589-595 (2009)
