[P006] Poster

1:00 PM - 1:50 PM JST(4:00 AM - 4:50 AM UTC)

[P03-356] Promoter-driven and translational tuning strategies enable enhanced aromatic compound biosynthesis in cyanobacterium Synechococcus sp. PCC 7002

○Hanh Duy Dao¹, Kenya Tanaka^1,2, Tomohisa Hasunuma^1,2,3 (1. Engineering Biology Research Center, Kobe University (Japan), 2. Graduate School of Science, Innovation and Technology, Kobe University (Japan), 3. Center for Sustainable Resource Science, RIKEN (Japan))

Keywords:

Cyanobacterial metabolic engineering,Aromatic compound biosynthesis,Promoter engineering,Ribosome binding site (RBS) engineering,CRISPRi

Aromatic compounds such as 4-hydroxybenzoate (4-HB) and phenylalanine (Phe) are valuable platform chemicals with broad industrial applications. However, their production in cyanobacteria is often constrained by limited precursor availability, metabolic burden due to overexpression gene and competing metabolic flux within the shikimate pathway. Here, we developed a multi-level regulatory engineering strategy in Synechococcus sp. PCC 7002 to enhance aromatic biosynthesis through coordinated transcriptional and translational control. A hybrid pathway was constructed by expressing the native chorismate pyruvate-lyase (UbiC) from Synechococcus sp. PCC 7002 together with heterologous feedback-resistant DAHP synthase (aroG) from Escherichia coli, while reinforcing native phosphoenolpyruvate synthase (ppsA), under the control of the native carbon-responsive sbtA promoter (from the sodium-dependent bicarbonate transporter gene sbtA) to produce 4-HB. This design enabled the direct conversion of chorismate to 4-HB, while the sbtA promoter served as an alternative strategy for inducing potentially toxic heterologous genes without the need for expensive chemical inducers. CRISPRi-mediated repression of aroH, encoding chorismate mutase that converts chorismate to prephenate in the phenylalanine biosynthesis pathway, was implemented to reduce competing carbon flux and redirect chorismate toward 4-HB, increasing its production from 70.5 mg/L to 185 mg/L.To enhance Phe production, the above construct lacking UbiC was further engineered by optimizing the ribosome binding site of aroG to fine-tune translation efficiency. Interestingly, the variant with the lowest predicted translation rate resulted in the highest Phe production (996 mg/L), highlighting the importance of translational balancing for optimal carbon distribution.The present study demonstrates that integrating promoter-driven transcriptional control with translational tuning is an effective strategy to optimize aromatic compound production in cyanobacteria. These findings advance Synechococcus sp. PCC 7002 as a sustainable photosynthetic platform for the biosynthesis of value-added aromatics.
[Acknowledgements]. This work was supported by GteX Program Japan (grant number JPMJGX23B4).