Summary The biotechnological production of succinate bears serious potential to fully replace existing petrochemical approaches in the future. In order to establish an economically viable bioprocess, obtaining high titre, yield and productivity is of central importance. In this study, we present a straightforward engineering approach for anaerobic succinate production with Vibrio natriegens, consisting of essential metabolic engineering and optimization of process conditions. The final producer strain V. natriegens Δlldh Δdldh Δpfl Δald Δdns::pycCg (Succ1) yielded 1.46 mol of succinate per mol of glucose under anaerobic conditions (85% of the theoretical maximum) and revealed a particularly high biomass‐specific succinate production rate of 1.33 gSucc gCDW−1 h−1 compared with well‐established production systems. By applying carbon and redox balancing, we determined the intracellular flux distribution and show that under the tested conditions the reductive TCA as well as the oxidative TCA/glyoxylate pathway contributed to succinate formation. In a zero‐growth bioprocess using minimal medium devoid of complex additives and expensive supplements, we obtained a final titre of 60.4 gSucc l−1 with a maximum productivity of 20.8 gSucc l−1 h−1 and an overall volumetric productivity of 8.6 gSucc l−1 h−1 during the 7 h fermentation. The key performance indicators (titre, yield and productivity) of this first engineering approach in V. natriegens are encouraging and compete with costly tailored microbial production systems.
Background Itaconic acid is a promising platform chemical for a bio-based polymer industry. Today, itaconic acid is biotechnologically produced with Aspergillus terreus at industrial scale from sugars. The production of fuels but also of chemicals from food substrates is a dilemma since future processes should rely on carbon sources which do not compete for food or feed. Therefore, the production of chemicals from alternative substrates such as acetate is desirable to develop novel value chains in the bioeconomy. Results In this study, Corynebacterium glutamicum ATCC 13032 was engineered to efficiently produce itaconic acid from the non-food substrate acetate. Therefore, we rewired the central carbon and nitrogen metabolism by inactivating the transcriptional regulator RamB, reducing the activity of isocitrate dehydrogenase, deletion of the gdh gene encoding glutamate dehydrogenase and overexpression of cis-aconitate decarboxylase (CAD) from A. terreus optimized for expression in C. glutamicum. The final strain C. glutamicum ΔramB Δgdh IDHR453C (pEKEx2-malEcadopt) produced 3.43 ± 0.59 g itaconic acid L−1 with a product yield of 81 ± 9 mmol mol−1 during small-scale cultivations in nitrogen-limited minimal medium containing acetate as sole carbon and energy source. Lowering the cultivation temperature from 30 °C to 25 °C improved CAD activity and further increased the titer and product yield to 5.01 ± 0.67 g L−1 and 116 ± 15 mmol mol−1, respectively. The latter corresponds to 35% of the theoretical maximum and so far represents the highest product yield for acetate-based itaconic acid production. Further, the optimized strain C. glutamicum ΔramB Δgdh IDHR453C (pEKEx2-malEcadopt), produced 3.38 ± 0.28 g itaconic acid L−1 at 25 °C from an acetate-containing aqueous side-stream of fast pyrolysis. Conclusion As shown in this study, acetate represents a suitable non-food carbon source for itaconic acid production with C. glutamicum. Tailoring the central carbon and nitrogen metabolism enabled the efficient production of itaconic acid from acetate and therefore this study offers useful design principles to genetically engineer C. glutamicum for other products from acetate.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.