Entwicklung der pflanzlichen Proteinumsatzrate zur Erhöhung der Biomasse
Biochemie
Zusammenfassung der Projektergebnisse
The world population is expected to exceed 9 billion by 2050, and overall production of the major food crops must raise at least 50% to ensure adequate nutrition for all these people. Yield improvement is already starting to plateau, so conventional breeding will likely not suffice to reach this goal. This makes development and application of new tools urgent to increase crop biomass and harvested yields. Metabolic engineering and synthetic biology are such tools. They can decrease carbon loss by lowering respiration rates, thereby saving carbon that can subsequently be channeled into biomass. At least half of the carbon fixed by photosynthesis can never be stored in biomass because it is consumed in respiration. Part of that fuels protein turnover which makes up to 20% of a crop’s total energy budget. Protein turnover (= degradation and resynthesis) is important to replace damaged enzymes, e.g. upon self-inactivation. Enzyme self-inactivation is a widely underestimated cause of protein turnover; however, enzymes which perform dangerous chemistry can easily undergo inactivation caused by the reaction catalyzed, e.g., by reactive substrates, products, or cofactors. Compared to long-lived enzymes which can catalyze millions of reactions before inactivation, short-lived enzymes catalyze a single or a couple thousand reactions. Cutting costs of abundant but short-lived enzymes is an almost totally unexplored strategy to minimize carbon loss and to engineer biomass productivity. It is a suitable target to reduce respiratory costs in crops. The funded study aimed at increasing the working lifetime in vivo of THI4 thiazole synthase and methionine synthase (MS). Both are exceptionally short-lived but abundant enzymes in plants. In an exploratory study we first accessed the engineering potential of Arabidopsis MS1. We found that vulnerable amino acids in or close to the active center are replaced by more stable amino acids in bacterial sequences. This was a good indication that there is potential to damage-harden plant enzymes. In a pioneer study, I then evolved plant THI4s and MS to live longer in vivo. For that, I used a toxic analogue strategy and the continuous directed evolution tool OrthoRep in yeast. Good progress was made in evolving THI4, and the project has recently been taken over by a coworker for further research. The MS was engineered in OrthoRep to overproduce methionine. Analytics showed that evolved populations produced up to 12-times more methionine – a strong indication for longer working life. Validation of mutant proteins in fresh OrthoRep cells, by in vitro assays and protein abundance measurements is in progress and will be finished for publication by June. In addition, we published a review paper about the respiratory energy demands and scope for demand expansion and destruction2. This paper was meant to estimate the costs and benefits of popular crop synthetic biology interventions, e.g. biological nitrogen fixation, cut respiration and carbon sequestration.
Projektbezogene Publikationen (Auswahl)
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The Moderately (D)efficient Enzyme: Catalysis-Related Damage In Vivo and Its Repair. Biochemistry, 60(47), 3555-3565.
Bathe, Ulschan; Leong, Bryan J.; McCarty, Donald R.; Henry, Christopher S.; Abraham, Paul E.; Wilson, Mark A. & Hanson, Andrew D.
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Trichomes: Tiny Plant Organs with Superpower. Specialty Crop Industry
U. Bathe
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Using continuous directed evolution to improve enzymes for plant applications. Plant Physiology, 188(2), 971-983.
García-García, Jorge D.; Van Gelder, Kristen; Joshi, Jaya; Bathe, Ulschan; Leong, Bryan J.; Bruner, Steven D.; Liu, Chang C. & Hanson, Andrew D.
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Blueberry: Behind the Superfood Status. Specialty Crop Industry
U. Bathe & B. Leong
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Fluoroacetate distribution, response to fluoridation, and synthesis in juvenile Gastrolobium bilobum plants. Phytochemistry, 202, 113356.
Leong, Bryan J.; Folz, Jacob S.; Bathe, Ulschan; Clark, David G.; Fiehn, Oliver & Hanson, Andrew D.
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Respiratory energy demands and scope for demand expansion and destruction. Plant Physiology, 191(4), 2093-2103.
Bathe, Ulschan; Leong, Bryan J.; Van Gelder, Kristen; Barbier, Guillaume G.; Henry, Christopher S.; Amthor, Jeffrey S. & Hanson, Andrew D.
