Presentation Information
[3GteX-11-KL]Systematic understanding and engineering of Calvin cycle in cyanobacteria
○Kenya Tanaka1,2,3, Akihiko Kondo2, Tomohisa Hasunuma1,2,4,5 (1. Engineering Biology Research Center, Kobe University (Japan), 2. Graduate School of Science, Innovation and Technology, Kobe University (Japan), 3. Research Center for Solar Energy Chemistry, Graduate School of Engineering Science, The University of Osaka (Japan), 4. RIKEN Center for Sustainable Resource Science (Japan), 5. Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University (Japan))
Keywords:
photosynthesis,metabolome,redox proteomics,metabolic engineering
Photosynthetic microorganisms, such as cyanobacteria, play a critical role in global carbon cycling through CO2 fixation, while exhibiting remarkable metabolic flexibility to cope with environmental fluctuations. To understand and engineer the robustness of these organisms, we must elucidate the dynamic and regulatory mechanisms that govern their metabolic networks during environmental transitions.
One such transition, from dark to light conditions, induces a rapid shift from heterotrophic to autotrophic metabolism, requiring a tightly coordinated activation of the Calvin–Benson–Bassham (CBB) cycle. We have systematically characterized this transition in Synechocystis sp. PCC 6803 by integrating absolute metabolite quantification with time-resolved 13C-tracer experiments. Our analysis revealed that glycolytic intermediates such as phosphoglycerate and phosphoenolpyruvate, which accumulate during dark adaptation, serve as essential substrates to jump-start CO2 fixation upon illumination (1). Moreover, overexpression of pyruvate kinase significantly impaired the initiation of photosynthesis, underscoring the importance of metabolic pre-conditioning for environmental robustness (2).
In parallel, we have explored another fundamental regulatory layer: redox-mediated enzyme control via reversible disulfide bond formation. To systematically map redox potentials of cysteine pairs across the proteome, we developed a novel method involving redox equilibration buffers, iodo-TMT labeling, and LC-MS/MS-based quantification. Application of this method to Synechocystis allowed estimation of in vitro redox potentials for a wide range of proteins, providing a new resource to dissect redox regulation in photosynthetic cells. These data offer insights into how redox signaling fine-tunes metabolism and complements metabolite-level regulation under fluctuating conditions.
Together, these studies demonstrate the power of combining metabolomics, isotope labeling, and redox proteomics to gain a systems-level understanding of how photosynthetic microorganisms maintain metabolic robustness. Such insights will be critical for future metabolic engineering efforts aimed at enhancing photosynthetic productivity under variable environments.
(1) Tanaka K, Shirai T, Vavricka CJ, Matsuda M, Kondo A, Hasunuma T, Plant Physiology, 191, 2400-2413 (2023)
(2) Tanaka K, Kondo A, Hasunuma T, Plant Cell Physiology, 65, 1812-1820 (2024)
One such transition, from dark to light conditions, induces a rapid shift from heterotrophic to autotrophic metabolism, requiring a tightly coordinated activation of the Calvin–Benson–Bassham (CBB) cycle. We have systematically characterized this transition in Synechocystis sp. PCC 6803 by integrating absolute metabolite quantification with time-resolved 13C-tracer experiments. Our analysis revealed that glycolytic intermediates such as phosphoglycerate and phosphoenolpyruvate, which accumulate during dark adaptation, serve as essential substrates to jump-start CO2 fixation upon illumination (1). Moreover, overexpression of pyruvate kinase significantly impaired the initiation of photosynthesis, underscoring the importance of metabolic pre-conditioning for environmental robustness (2).
In parallel, we have explored another fundamental regulatory layer: redox-mediated enzyme control via reversible disulfide bond formation. To systematically map redox potentials of cysteine pairs across the proteome, we developed a novel method involving redox equilibration buffers, iodo-TMT labeling, and LC-MS/MS-based quantification. Application of this method to Synechocystis allowed estimation of in vitro redox potentials for a wide range of proteins, providing a new resource to dissect redox regulation in photosynthetic cells. These data offer insights into how redox signaling fine-tunes metabolism and complements metabolite-level regulation under fluctuating conditions.
Together, these studies demonstrate the power of combining metabolomics, isotope labeling, and redox proteomics to gain a systems-level understanding of how photosynthetic microorganisms maintain metabolic robustness. Such insights will be critical for future metabolic engineering efforts aimed at enhancing photosynthetic productivity under variable environments.
(1) Tanaka K, Shirai T, Vavricka CJ, Matsuda M, Kondo A, Hasunuma T, Plant Physiology, 191, 2400-2413 (2023)
(2) Tanaka K, Kondo A, Hasunuma T, Plant Cell Physiology, 65, 1812-1820 (2024)
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