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Refining and illuminating acetogenic Eubacterium strains for reclassification and metabolic engineering
Maximilian Flaiz1,2, Anja Poehlein3, Wiebke Wilhelm4
1Laboratory of Microbiology, Wageningen University and Research, Wageningen, The Netherlands. maximilian.flaiz@wur.nl.
Microbial Cell Factories
|January 17, 2024
Summary
Eubacterium strains, including E. limosum, are excellent biocatalysts for converting C1 substrates into valuable products like butyrate. Researchers successfully genetically engineered eleven strains, enabling new biotechnological applications.
Area of Science:
- Microbiology
- Synthetic Biology
- Biotechnology
Background:
- The genus Eubacterium contains diverse acetogenic strains capable of fermenting C1 substrates.
- Eubacterium limosum and related species are notable for C1 gas/liquid fermentation and butyrate production.
- E. limosum's well-understood metabolism and genetic accessibility make it a promising industrial biocatalyst.
Purpose of the Study:
- To investigate the genomic, phylogenetic, and physiological characteristics of E. limosum and its close relatives.
- To identify genetic elements responsible for C1 substrate utilization and chain elongation.
- To establish a reliable genetic engineering protocol for industrial applications.
Main Methods:
- Genome sequencing and comparative analysis of eleven Eubacterium strains.
- Phylogenetic analysis to establish distinct clades (E. limosum, E. callanderi, E. maltosivorans).
- Growth experiments to assess methanol conversion and product formation (acetate, butyrate, hexanoate).
- Development and application of a harmonized electroporation protocol for genetic transformation.
- Utilizing the fluorescence-activating and absorption shifting tag (FAST) as a reporter for genetic engineering verification via flow cytometry.
Main Results:
- Eleven Eubacterium strains were classified into three distinct clades: E. limosum, E. callanderi, and E. maltosivorans.
- Identified gene clusters for methanol utilization and chain elongation in all analyzed strains.
- Demonstrated that strains from all three clades can convert methanol to acetate, butyrate, and hexanoate via reverse β-oxidation.
- Successfully genetically engineered eight Eubacterium strains using a harmonized electroporation protocol and FAST reporter.
Conclusions:
- E. limosum, E. callanderi, and E. maltosivorans are prime candidates for anaerobic C1-substrate bioconversion.
- A standardized electroporation protocol enables genetic engineering across multiple Eubacterium strains.
- FAST serves as an effective fluorescent reporter for characterizing engineered Eubacterium.
- Updated classification of eleven strains into distinct clades, with proposed emendations to species descriptions for database implementation.
Keywords:
AcetogensAnaerobesEubacteriumFASTFluorescenceHexanoateMethanolcallanderilimosummaltosivoransMore Related Videos
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