Engineering of itaconic acid pathway via co-localization of CadA and AcnA in recombinant Escherichia coli

Itaconic acid is an excellent polymeric precursor with a wide range of industrial applications. The efficient production of itaconate from various renewable substrates was demonstrated by engineered Escherichia coli. However, limitation in the itaconic acid precursor supply was revealed by finding out the key intermediate of the tricarboxylic acid in the itaconic acid pathway. Efforts of enhancing the cis-aconitate flux and preserving the isocitrate pool to increase itaconic acid productivity are required. In this study, we introduce a synthetic protein scaffold system between CadA and AcnA to physically combine the two enzymes. Through the introduction of a synthetic protein scaffold, 2.1 g L^-1 of itaconic acid was produced at pH 7 and 37 °C. By fermentation, 20.1 g L^-1 for 48 h of itaconic acid was produced with a yield of 0.34 g g^-1 glycerol. These results suggest that carbon flux was successfully increased itaconic acid productivity.