Metabolic engineering of Escherichia coli to produce (2S, 3R, 4S)-4-hydroxyisoleucine |
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Authors: | Sergey V Smirnov Tomohiro Kodera Natalya N Samsonova Veronika А Kotlyarovа Natalya Yu Rushkevich Аlexander D Kivero Pavel M Sokolov Makoto Hibi Jun Ogawa Sakayu Shimizu |
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Institution: | 1. Ajinomoto-Genetika Research Institute, 1st Dorozhny pr. 1, Moscow, 117545, Russia 2. Institute of Life Sciences, Ajinomoto Co., Inc., Suzuki-cho, Kawasaki-ku, Kawasaki-shi, 210-8681, Japan 3. Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, 8 Sakyo-ku, Kyoto, 606-8502, Japan
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Abstract: | The stereo-specific l-isoleucine-4-hydroxylase (l-isoleucine dioxygenase (IDO)) was cloned and expressed in an Escherichia coli 2Δ strain lacking the activities of α-ketoglutarate dehydrogenase (EC 1.2.4.2), isocitrate liase (EC 4.1.3.1), and isocitrate dehydrogenase kinase/phosphatase (EC 2.7.11.5). The 2Δ strain could not grow in a minimal-salt/glucose/glycerol medium due to the blockage of TCA during succinate synthesis. The IDO activity in the 2Δ strain was able to “shunt” destroyed TCA, thereby coupling l-isoleucine hydroxylation and cell growth. Using this strain, we performed the direct biotransformation of l-isoleucine into 4-HIL with an 82% yield. |
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