Xixian Chen / Agency for Science, Technology and Research, Singapore, Singapore
Rehka T / Agency for Science, Technology and Research, Singapore, Singapore
Jeremy Esque / Toulouse biotechnology Institute, Toulouse, France
Congqiang Zhang / Agency for Science, Technology and Research, Singapore, Singapore
Isabelle André / Toulouse biotechnology Institute, Toulouse, France
Topic: Synthetic Biology and Metabolic Engineering
Metabolic engineering is an attractive method for the efficient production of natural products. However, one critical pre-requisite is to establish the biosynthetic pathways. Many commercially interesting molecules cannot be biosynthesized as their native pathways are not fully elucidated. Cis-α-irone, a top-end perfumery molecule, is an example. To address this challenge, retrobiosynthetic pathway design with promiscuous enzymes provides an alternative solution. Here, we design a synthetic pathway to produce cis-α-irone with a promiscuous methyltransferase (pMT). Using structure-guided enzyme engineering strategies, we improved pMT activity by >10,000-fold and specificity by >1000-fold. 120 mg/L cis- α-irone is produced from psi-ionone. 86 mg/L cis-α-irone is produced from glucose by engineered microbe in a 5 l bioreactor. Our work illustrates that integrated retropathway design and enzyme engineering can offer opportunities to expand the scope of molecules that can be biosynthesized.