P178: Engineering a novel biosynthetic pathway for rosmarinic acid in Escherichia coli

Phenylpropanoids are plant secondary metabolites with a broad range of structures and biological activities.  Due to the importance of these compounds in industry and pharmacy, there is a need for inexpensive, efficient means of production.  We aim to establish a microbial platform for the production of the hydroxycinnamate ester rosmarinic acid (RA).  RA is an antioxidant found in many culinary herbs and is used as a natural food preservative.  It is also a GABA-transaminase inhibitor, giving it potential as a drug in the treatment of anxiety or related disorders.  In plants, the RA biosynthetic pathway requires ten enzymatic steps, including catalysis by cytochrome P450s (CYPs), requiring CYP reductases which are absent in most industrial microbes.  By seeking out bacterial enzymes shown to have the same or similar activities to the plant enzymes, we have designed a novel route to RA biosynthesis in Escherichia coli requiring only six enzymes, four of which are bacterial in origin.  We have assembled subsets of the pathway and shown production of RA precursors.  The final pathway enzyme, RA synthase, has been shown to accept a range of substrates to synthesize a number of hydroxycinnamate esters and amides.  We aim to use the bacterial RA pathway to produce novel hydroxycinnamate esters through combinatorial and in vitro evolution approaches.