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200102

Project description:

Project no:
200102
Title:
Development of high-efficiency Cupriavidus necator strains for the production of platform chemicals from carbon-dioxide
1st project leader:
Gescher, Johannes - Institute for Applied Biology (IAB) Department of Applied Biology, Karlsruhe Institute of Technology (KIT), Karlsruhe
2nd project leader
Description:
Motivation: The development of a bioeconomy needs processes for the biotechnological conversion of carbon dioxide to platform chemicals. So far, the spectrum of typical biocatalysts for a biotechnological conversion of carbon dioxide to valuable chemicals is rather small and typically includes methanogens and acetogens. These organisms produce a narrow spectrum of end-products and are difficult to modify genetically. Recently, we developed a Cupriavidus necator strain that produces platform chemicals with a very high carbon efficiency(1). The organism belongs to the group of knallgas bacteria and thrives with hydrogen, CO2 and oxygen. Although the carbon efficiency of the organism is almost 100% the electron efficiency is lower compared to methanogens and acetogens most probably due to the unspecifity of the CO2-fixing enzyme Ribulose-bisphosphat carboxylase/oxygenase (RuBisCo) that also reduces oxygen in a side reaction.

Aim: The aim of the project is to raise the efficacy of the strain by increasing the intracellular carbon dioxide concentration and decrease the intracellular concentration of oxygen in the vicinity of RuBisCo.

Solution: The solution will be to introduce synthetic microcompartments called carboxysomes into the organism. Recently, the possibility to heterologously produce these microcompartments was shown in another microorganism(2). Microorganisms localize in these carboxysomes specifically two enzymes, the RubisCo and a carboanhydrase which increases the availability of carbon dioxide. Moreover, it was shown that a specific adjustment of the different carboanhydrases of the organism could potentially lead to a higher carbon dioxide availability. Hence, in parallel to the formation of synthetic microcompartments, it is the further aim to adjust the transcription level of the carboanhydrases to an optimum.
References:
1. Windhorst, C. & Gescher, J. Efficient biochemical production of acetoin from carbon dioxide using
Cupriavidus necator H16. Biotechnol Biofuels 12, 163–11 (2019).

2. Bonacci, W. et al. Modularity of a carbon-fixing protein organelle. Proc. Natl. Acad. Sci. U.S.A. 109, 478–483 (2012).

3. Gai, C. S., Lu, J., Brigham, C. J., Bernardi, A. C. & Sinskey, A. J. Insights into bacterial CO2 metabolism revealed by the characterization of four carbonic anhydrases in Ralstonia eutropha H16. AMB Express 4, 2(2014).
Methods that will be used:
Genetic rearrangements will all be conducted and analyzed by state-of-the-art methods (e.g. Gibson cloning, transcriptomics). Imaging techniques will be used to identify the microcompartments inside the C. necator strains. Micro-GC, HPLC and quantitative PCR experiments will be conducted to assess the efficiency of the organism with regards to the demand of electron per carbon dioxide molecule reduced.
Collaboration partners:
Prof. Harald Horn, KIT, Karlsruhe
Expected candidate‘s qualification:
Good English language skills, basic knowledge on microbial physiology, practical knowledge on molecular biology tools for genetic engineering in prokaryotes.
Keywords:
Gas fermentation, synthetic biology, Cupriavidus