Model-based metabolic engineering enables high yield itaconic acid production by Escherichia coli.
Itaconic acid is a high potential platform chemical which is currently industrially produced by Aspergillus terreus. Heterologous production of itaconic acid with Escherichia coli could help to overcome limitations of A. terreus regarding slow growth and high sensitivity to oxygen supply. However, the performance achieved so far with E. coli strains is still low. We introduced a plasmid (pCadCS) carrying genes for itaconic acid production into E. coli and applied a model-based approach to construct a high yield production strain. Based on the concept of minimal cut sets, we identified intervention strategies that guarantee high itaconic acid yield while still allowing growth. One cut set was selected and the corresponding genes were iteratively knocked-out. As a conceptual novelty, we pursued an adaptive approach allowing changes in the model and initially calculated intervention strategy if a genetic modification induces changes in byproduct formation. Using this approach, we iteratively implemented five interventions leading to high yield itaconic acid production in minimal medium with glucose as substrate supplemented with small amounts of glutamic acid. The derived E. coli strain (ita23: MG1655 ∆aceA ∆sucCD ∆pykA ∆pykF ∆pta ∆Picd::cam_BBa_J23115 pCadCS) synthesized 2.27g/l itaconic acid with an excellent yield of 0.77mol/(mol glucose). In a fed-batch cultivation, this strain produced 32g/l itaconic acid with an overall yield of 0.68mol/(mol glucose) and a peak productivity of 0.45g/l/h. These values are by far the highest that have ever been achieved for heterologous itaconic acid production and indicate that realistic applications come into reach.
Max Planck Institute for Dynamics of Complex Technical Systems, Sandtorstraße 1, 39106 Magdeburg, Germany.
Björn-Johannes
BJ
Harder
Max Planck Institute for Dynamics of Complex Technical Systems, Sandtorstraße 1, 39106 Magdeburg, Germany.
Katja
K
Bettenbrock
Max Planck Institute for Dynamics of Complex Technical Systems, Sandtorstraße 1, 39106 Magdeburg, Germany. Electronic address: klamt@mpi-magdeburg.mpg.de.
Steffen
S
Klamt
D053898:Biosynthetic Pathways / Q000502:physiology; D003198:Computer Simulation; D004926:Escherichia coli / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D024861:Genetic Enhancement / Q000379:methods*; D060847:Metabolic Engineering / Q000379:methods*; D064688:Metabolic Flux Analysis / Q000379:methods*; D053858:Metabolic Networks and Pathways / Q000502:physiology; D008954:Models, Biological; D015398:Signal Transduction / Q000502:physiology; D013386:Succinates / Q000302:isolation & purification / Q000378:metabolism
D029968:Escherichia coli Proteins; D013386:Succinates; C005229:itaconic acid
Biotechnology; Computational strain design; Constraint-based and stoichiometric modeling; Escherichia coli; Itaconate production; Itaconic acid (PubChem CID: 811); Metabolic engineering; Microbial cell factories