Improvement of dicarboxylic acid production with Methylorubrum extorquens by reduction of product reuptake

  • The methylotrophic bacterium Methylorubrum extorquens AM1 has the potential to become a platform organism for methanol-driven biotechnology. Its ethylmalonyl-CoA pathway (EMCP) is essential during growth on C1 compounds and harbors several CoA-activated dicarboxylic acids. Those acids could serve as precursor molecules for various polymers. In the past, two dicarboxylic acid products, namely mesaconic acid and 2-methylsuccinic acid, were successfully produced with heterologous thioesterase YciA from Escherichia coli, but the yield was reduced by product reuptake. In our study, we conducted extensive research on the uptake mechanism of those dicarboxylic acid products. By using 2,2-difluorosuccinic acid as a selection agent, we isolated a dicarboxylic acid import mutant. Analysis of the genome of this strain revealed a deletion in gene dctA2, which probably encodes an acid transporter. By testing additional single, double, and triple deletions, we were able to rule out the involvement of the two other DctA transporter homologs and the ketoglutarate transporter KgtP. Uptake of 2-methylsuccinic acid was significantly reduced in dctA2 mutants, while the uptake of mesaconic acid was completely prevented. Moreover, we demonstrated M. extorquens-based synthesis of citramalic acid and a further 1.4-fold increase in product yield using a transport-deficient strain. This work represents an important step towards the development of robust M. extorquens AM1 production strains for dicarboxylic acids.

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Author:Laura PöschelORCiDGND, Elisabeth Gehr, Markus BuchhauptORCiDGND
Parent Title (English):Applied microbiology and biotechnology
Place of publication:Berlin ; Heidelberg ; New York
Document Type:Article
Date of Publication (online):2022/09/15
Date of first Publication:2022/09/15
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2023/11/13
Tag:Acid transporters; Dicarboxylic acids; Ethylmalonyl-CoA; Methylorubrum extorquens; Methylotroph; Product reuptake; pathway
Page Number:19
First Page:6713
Last Page:6731
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie
Licence (German):License LogoCreative Commons - CC BY - Namensnennung 4.0 International