%0 Journal Article 
%A Valle Gallardo, Antonio
%A Soto, Zamira
%A Muhamadali, Howbeer
%A Hollywood, Katherine A.
%A Xu, Yun
%A Lloyd, Jonathan R.
%A Goodacre, Royston
%A Cantero Moreno, Domingo
%A Cabrera Revuelta, Gema
%A Bolívar Pérez, Jorge
%T Metabolomics for the design of new metabolic engineering strategies for improving aerobic succinic acid production in Escherichia coli
%D 2022
%@ 1573-3882
%U http://hdl.handle.net/10498/27350
%X Introduction Glycerol is a byproduct from the biodiesel industry that can be biotransformed by Escherichia coli to high added-value products such as succinate under aerobic conditions. The main genetic engineering strategies to achieve this aim involve the mutation of succinate dehydrogenase (sdhA) gene and also those responsible for acetate synthesis including acetate kinase, phosphate acetyl transferase and pyruvate oxidase encoded by ackA, pta and pox genes respectively in the Delta sdhA Delta ack-pta Delta pox (M4) mutant. Other genetic manipulations to rewire the metabolism toward succinate consist on the activation of the glyoxylate shunt or blockage the pentose phosphate pathway (PPP) by deletion of isocitrate lyase repressor (iclR) or gluconate dehydrogenase (gnd) genes on M4-Delta iclR and M4-Delta gnd mutants respectively.

Objective To deeply understand the effect of the blocking of the pentose phosphate pathway (PPP) or the activation of the glyoxylate shunt, metabolite profiles were analyzed on M4-Delta gnd, M4-Delta iclR and M4 mutants.

Methods Metabolomics was performed by FT-IR and GC-MS for metabolite fingerprinting and HPLC for quantification of succinate and glycerol.

Results Most of the 65 identified metabolites showed lower relative levels in the M4-Delta iclR and M4-Delta gnd mutants than those of the M4. However, fructose 1,6-biphosphate, trehalose, isovaleric acid and mannitol relative concentrations were increased in M4-Delta iclR and M4-Delta gnd mutants. To further improve succinate production, the synthesis of mannitol was suppressed by deletion of mannitol dehydrogenase (mtlD) on M4-Delta gnd Delta mtlD mutant that increase similar to 20% respect to M4-Delta gnd.

Conclusion Metabolomics can serve as a holistic tool to identify bottlenecks in metabolic pathways by a non-rational design. Genetic manipulation to release these restrictions could increase the production of succinate.
%K Escherichia coli
%K Metabolomics
%K Succinic acid
%K TCA cycle
%K GC–MS and mannitol dehydrogenase (MtlD)
%~ Universidad de Cádiz