Growth of Escherichia coli on protocatechuic acid via a chromosomally integrated synthetic ortho-degradation pathway

Lignocellulosic material, such as wood, grass, and agricultural and municipal waste, is a potential renewable source of carbon for production of chemicals. Creating value-added products from lignin will increase the productivity and profitability of using lignocellulosic materials for chemical generation. Escherichia coli is a commonly engineered platform organism for the biological production of fuels and chemicals, but it is incapable of converting many phenolic compounds such as protocatechuate (PCA). The β-ketoadipate pathway funnels carbon from a variety of aromatic compounds to tricarboxylic acid cycle intermediates, with one branch consisting of the PCA ortho-degradation pathway. In this work, we introduced into E. coli the PCA ortho-degradation pathway from Pseudomonas putida KT2440, as well as the PCA transporter, pcaK. Heterologous expression of this pathway from high-copy plasmids permitted growth using PCA as the sole carbon and energy source. However, growth of this strain was poor on both glucose and on PCA, suggesting inhibition derived from overproduction of the PCA degradation pathway. Integration of this pathway into the chromosome resulted in faster growth on PCA with no defect during growth on glucose. This PCA degrading E. coli strain can serve as a platform for further engineering to convert lignin-derived compounds into value-added chemicals by E. coli.