Metabolic rewiring of Synechococcus elongatus PCC 7942 enhances astaxanthin production from CO2

Photosynthetic cyanobacteria are promising chassis for converting CO2 into value-added products, although competition between cellular growth and biosynthesis can constrain productivity. Here, the high-carbon-fixing, adaptively evolved Synechococcus elongatus PCC 7942 strain S20 was engineered for astaxanthin production through coordinated regulation of precursor supply, pathway topology, and cofactor engineering. Heterologous expression of β-carotene ketolase and hydroxylase generated strain S20-5, which accumulated 7.04 mg/g dry cell weight (DCW) astaxanthin. Two strategies were adopted to enhance astaxanthin production. First, precursor supply was strengthened by enhancing the endogenous methylerythritol phosphate pathway or introducing a heterologous isopentenol utilization pathway. These modifications promoted zeaxanthin and canthaxanthin accumulation without further increasing astaxanthin content, indicating that precursor supply alone was insufficient. Accordingly, the synthetic operon was reorganized to alter the transcriptional patterns of the ketolase and hydroxylase genes, generating strain S20-6, which accumulated 11.04 mg/g DCW astaxanthin, the highest content reported in S. elongatus PCC 7942 to date, with astaxanthin as the predominant carotenoid product. Second, lipoate-protein ligase A (LplA) was heterologously expressed as a cofactor-engineering strategy intended to reinforce lipoate-dependent central metabolic functions, generating strains S20-53 and S20-61. Under 10% CO2, the OD750 values of S20-53 and S20-61 increased by 81% and 39% relative to S20-5 and S20-6, respectively. These phenotypes position S20-6 as a candidate for product-oriented astaxanthin production and S20-53 as a platform for value-added biomanufacturing using high-concentration CO2. This study highlights the importance of coordinating precursor supply, downstream conversion, and host metabolic homeostasis when constructing efficient photosynthetic cell factories.