Co-Culture Based Modular Engineering for Aromatic and Aromatic-Derived Compounds Production in E. coli

Biosynthesis of aromatic and aromatic-derived compounds in engineered microbe provides a robust means to produce value-added molecules from inexpensive and renewable carbon sources. Here, we present a novel co-culture strategy to biosynthesize aromatic and aromatic-derived compounds using engineered E. coli. As a proof of concept, we show that production of industrially important compound muconic acid can be achieved in E. coli at high titer and yield. Specifically, the shikimate pathway of E. coli was first engineered to produce the key intermediate, dehydroshikimate (DHS), as a starting molecule for aromatics biosynthesis. The downstream heterologous biosynthetic pathway and a DHS transporter were then engineered in E. coli to produce muconic acid at a titer of 500 mg/L in test tube. Next, the whole pathway was split into two modules, each of which was carried by an individually engineered E. coli cell. The upstream cell was responsible for DHS production, whereas the downstream cell was dedicated to importing DHS to produce muconic acid. Such a modular engineering strategy reduced the metabolic stress on each cell and resulted in a 40% production improvement, representing 12% of theoretical maximum yield. Furthermore, the two-cell system was optimized so that each cell grew on different sugars, which enabled the co-culture to simultaneously consume glucose and xylose, the major two components of lignocellulose, to biosynthesize muconic acid without the glucose repression effect. The proposed strategy is readily applicable for microbial production of a variety of valuable aromatics or aromatic-derived compounds from inexpensive and renewable carbon sources.