Screening and Engineering the Synthetic Potential of Carboxylating Reductases from Central Metabolism and Polyketide Biosynthesis |
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Authors: | Dominik M Peter Lennart Schada von Borzyskowski Dr Patrick Kiefer Philipp Christen Prof Julia A Vorholt Dr Tobias J Erb |
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Institution: | 1. Biochemistry and Synthetic Biology of Microbial Metabolism Group, Max‐Planck‐Institute for terrestrial Microbiology, Karl‐von‐Frisch‐Strasse 10, 35043 Marburg (Germany);2. Institute of Microbiology, Eidgen?ssisch Technische Hochschule (ETH) Zürich, Vladimir‐Prelog‐Weg 4, 8050 Zürich (Switzerland) |
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Abstract: | Carboxylating enoyl‐thioester reductases (ECRs) are a recently discovered class of enzymes. They catalyze the highly efficient addition of CO2 to the double bond of α,β‐unsaturated CoA‐thioesters and serve two biological functions. In primary metabolism of many bacteria they produce ethylmalonyl‐CoA during assimilation of the central metabolite acetyl‐CoA. In secondary metabolism they provide distinct α‐carboxyl‐acyl‐thioesters to vary the backbone of numerous polyketide natural products. Different ECRs were systematically assessed with a diverse library of potential substrates. We identified three active site residues that distinguish ECRs restricted to C4 and C5‐enoyl‐CoAs from highly promiscuous ECRs and successfully engineered a selected ECR as proof‐of‐principle. This study defines the molecular basis of ECR reactivity, allowing for predicting and manipulating a key reaction in natural product diversification. |
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Keywords: | biotechnology CO2 fixation crotonyl‐CoA carboxylase/reductase natural product engineering polyketides |
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