{"id":177575,"date":"2026-06-11T07:26:00","date_gmt":"2026-06-11T05:26:00","guid":{"rendered":"https:\/\/renewable-carbon.eu\/news\/?p=177575"},"modified":"2026-06-03T16:51:51","modified_gmt":"2026-06-03T14:51:51","slug":"cyanobacterial-carbonic-anhydrases-drive-carbon-recycling-and-prodrug-biosynthesis-in-escherichia-coli","status":"publish","type":"post","link":"https:\/\/renewable-carbon.eu\/news\/cyanobacterial-carbonic-anhydrases-drive-carbon-recycling-and-prodrug-biosynthesis-in-escherichia-coli\/","title":{"rendered":"Cyanobacterial carbonic anhydrases drive carbon recycling and prodrug biosynthesis in\u00a0Escherichia coli"},"content":{"rendered":"\n\n\n

This study establishes a synthetic carbon dioxide-recycling platform in\u00a0Escherichia coli<\/em>\u00a0by integrating carbonic anhydrases from\u00a0Cyanobacterium aponinum<\/em>\u00a0together with cyanobacterial phosphoribulokinase and ribulose-1,5-bisphosphate carboxylase\/oxygenase (i.e., RuBisCO\u2013Prk). Expression of \u03b2-carbonic anhydrase (cca<\/em>A) and \u03b3- carbonic anhydrase-like scaffold proteins (ccm<\/em>M and\u00a0ccm<\/em>N) was decoupled to preserve enzyme accessibility and metabolic flexibility, while ribulose-1,5-bisphosphate carboxylase\/oxygenase\u2013prk<\/em>\u00a0modules provided the downstream carboxylation mechanism necessary for reassimilation of intracellular carbon dioxide. Functional validation through enzyme assays, gas-trap quantification, isotope tracing, and carbon flux modeling confirmed pathway activity and metabolic resilience at flask scale, with genetic stability supported by plasmid pairing and coexpression strategies. Finally, by focusing on the catalytic interplay between a triad of carbonic anhydrases,\u00a0prk<\/em>, and ribulose-1,5-bisphosphate carboxylase\/oxygenase, the system enables direct metabolic interfacing with biosynthetic pathways such as5-aminolevulinic acid, establishing a versatile framework for re-engineering microbial cell factories. At present, the platform operates at technology readiness level 3, representing a validated proof of concept at laboratory scale. Advancing beyond technology readiness level 3 will require mitigation of byproduct accumulation, expansion of host applicability beyond\u00a0Escherichia coli<\/em>, and demonstration of robustness under pilot-scale conditions. Addressing these challenges will be critical for progressing toward technology readiness level 4\u20135, where reproducibility and scalability can be systematically evaluated. This approach offers a complementary alternative to adaptive laboratory evolution and lays the foundation for climate-resilient biomanufacturing platforms relevant to sustainable chemical production.<\/strong><\/p>\n\n\n

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\"Graphical
Graphical abstract \u00a9 National Cheng Kung University<\/figcaption><\/figure><\/div>\n\n\n

Highlights<\/h3>\n\n\n\n

Cyanobacterial carbonic anhydrases, integrated with Calvin cycle modules, enable carbon dioxide recycling in Escherichia coli<\/em>, advancing minimalist organelle engineering.<\/p>\n\n\n\n

CRISPR interference -mediated repression of the native can<\/em> gene facilitated host adaptation for more effective carbon dioxide recycling during 5-aminolevulinic acid production.<\/p>\n\n\n\n

Coexpression of triad carbonic anhydrases along with ribulose-1,5-bisphosphate carboxylase\/oxygenase\u2013Phosphoribulokinase (RuBisCO\u2013Prk) genes in a can-deficient<\/em> strain led to a 1.46-fold gain in 5-aminolevulinic acid titer, along with a 39.4% reduction in carbon dioxide release.<\/p>\n\n\n\n

This engineered design demonstrates the potential of modular carbon dioxide recycling strategies in microbial cell factories, though broader benchmarking and translational framing would strengthen its positioning in the field.<\/p>\n\n\n\n

Abstract<\/h3>\n\n\n\n

Sustainable biomanufacturing in Escherichia coli<\/em> is advancing through the integration of carbon dioxide (CO2<\/sub>)-fixing modules from algal systems. However, the assimilation efficiency remains limited by ribulose-1,5-bisphosphate carboxylase\/oxygenase (RuBisCO) performance and intracellular CO2<\/sub> availability. To address these constraints, we introduced carbonic anhydrases from Cyanobacteria aponinum<\/em> (capo<\/em>CAs) and repressed the native CA (can<\/em>) gene using Clustered Regularly Interspaced Short Palindromic Repeats interference (CRISPRi) to enhance design specificity. Afterward, capo<\/em>CAs were overexpressed in E. coli<\/em> BL21(DE3) and its derivative strains of can<\/em> deficient (dCA), Calvin\u2013Benson\u2013Bassham (CBB) (RuBisCO\u2013Prk expressing), and dCA+CBB (coupled with dCA and CBB). For functional demonstration, the engineered strains were used to produce a valuable prodrug, 5-aminolevulinic acid (5-ALA). As a result, capo<\/em>CAs modestly improved 5-ALA titers, with further increments upon CBB coexpression. The optimum dCA+CBB* strain achieved 7.04 g\/l (1.46-fold improvement), alongside 9.59% carbon enrichment and a 39.4% reduction in CO2<\/sub> release. The 13<\/sup>C-isotope confirmed CO2<\/sub> incorporation during 5-ALA biosynthesis. This work provides tunable carbon-management modules for developing low-footprint microbial cell factories.<\/p>\n","protected":false},"excerpt":{"rendered":"

This study establishes a synthetic carbon dioxide-recycling platform in\u00a0Escherichia coli\u00a0by integrating carbonic anhydrases from\u00a0Cyanobacterium aponinum\u00a0together with cyanobacterial phosphoribulokinase and ribulose-1,5-bisphosphate carboxylase\/oxygenase (i.e., RuBisCO\u2013Prk). Expression of \u03b2-carbonic anhydrase (ccaA) and \u03b3- carbonic anhydrase-like scaffold proteins (ccmM and\u00a0ccmN) was decoupled to preserve enzyme accessibility and metabolic flexibility, while ribulose-1,5-bisphosphate carboxylase\/oxygenase\u2013prk\u00a0modules provided the downstream carboxylation mechanism necessary for […]<\/p>\n","protected":false},"author":114,"featured_media":0,"comment_status":"closed","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":{"_seopress_robots_primary_cat":"none","nova_meta_subtitle":"Cyanobacterial carbonic anhydrases, integrated with Calvin cycle modules, enable carbon dioxide recycling in\u00a0Escherichia coli, advancing minimalist organelle engineering","footnotes":""},"categories":[17143],"tags":[13383,5838,10416,21295,10453],"supplier":[3742],"class_list":["post-177575","post","type-post","status-publish","format-standard","hentry","category-recycling","tag-bacteria","tag-bioeconomy","tag-circulareconomy","tag-cyanobacteria","tag-recycling","supplier-national-cheng-kung-university"],"_links":{"self":[{"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/posts\/177575","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/users\/114"}],"replies":[{"embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/comments?post=177575"}],"version-history":[{"count":1,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/posts\/177575\/revisions"}],"predecessor-version":[{"id":177578,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/posts\/177575\/revisions\/177578"}],"wp:attachment":[{"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/media?parent=177575"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/categories?post=177575"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/tags?post=177575"},{"taxonomy":"supplier","embeddable":true,"href":"https:\/\/renewable-carbon.eu\/news\/wp-json\/wp\/v2\/supplier?post=177575"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}