caa a22 4500 605500746 CHVBK 20210128100557.0 cr unu---uuuuu 210128e20150601xx s 000 0 eng 10.1007/s00253-014-6308-0 doi (NATIONALLICENCE)springer-10.1007/s00253-014-6308-0 Functional expression of l -lysine α-oxidase from Scomber japonicus in Escherichia coli for one-pot synthesis of l -pipecolic acid from dl -lysine [Elektronische Daten] [Yasushi Tani, Ryoma Miyake, Ryoichi Yukami, Yasumasa Dekishima, Hideyasu China, Shigeki Saito, Hiroshi Kawabata, Hisaaki Mihara] l-Pipecolic acid is a key component of biologically active molecules and a pharmaceutically important chiral building block. It can be stereoselectively produced from l-lysine by a two-step bioconversion involving l-lysine α-oxidase and ∆1-piperideine-2-carboxylae (Pip2C) reductase. In this study, we focused on an l-lysine α-oxidase from Scomber japonicus that was originally identified as an apoptosis-inducing protein (AIP) and applied the enzyme to one-pot fermentation of l-pipecolic acid in Escherichia coli. A synthetic gene coding for an AIP was expressed in E. coli, and the recombinant enzyme was purified and characterized. The purified enzyme was determined to be a homodimer with a molecular mass of 133.9kDa. The enzyme essentially exhibited the same substrate specificity as the native enzyme. Optimal temperature and pH for the enzymatic reaction were 70 °Cand 7.4, respectively. The enzyme was stable below 60°C and at a pH range of 5.5-7.5 but was markedly inhibited by Co2+. To establish a one-pot fermentation system for the synthesis of optically pure l-pipecolic acid from dl-lysine, an E. coli strain carrying a plasmid encoding AIP, Pip2C reductase from Pseudomonas putida, lysine racemase from P. putida, and glucose dehydrogenase from Bacillus subtilis was constructed. The one-pot process produced 45.1g/L of l-pipecolic acid (87.4% yield from dl-lysine) after a 46-h reaction with high optical purity (>99.9% enantiomeric excess). Springer-Verlag Berlin Heidelberg, 2014 l- Lysine α-oxidase nationallicence l- Pipecolic acid nationallicence One-pot synthesis nationallicence Stereoselective synthesis nationallicence ∆1-Piperideine-2-carboxylic acid nationallicence Tani Yasushi Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut Miyake Ryoma Mitsubishi Chemical Group Science and Technology Research Center, Inc., 227-8502, Yokohama, Japan aut Yukami Ryoichi Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut Dekishima Yasumasa Mitsubishi Chemical Group Science and Technology Research Center, Inc., 227-8502, Yokohama, Japan aut China Hideyasu Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut Saito Shigeki Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut Kawabata Hiroshi Mitsubishi Chemical Group Science and Technology Research Center, Inc., 227-8502, Yokohama, Japan aut Mihara Hisaaki Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/12(2015-06-01), 5045-5054 0175-7598 99:12<5045 2015 99 253 https://doi.org/10.1007/s00253-014-6308-0 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-014-6308-0 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Tani Yasushi Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut NATIONALLICENCE 700 1- Miyake Ryoma Mitsubishi Chemical Group Science and Technology Research Center, Inc., 227-8502, Yokohama, Japan aut NATIONALLICENCE 700 1- Yukami Ryoichi Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut NATIONALLICENCE 700 1- Dekishima Yasumasa Mitsubishi Chemical Group Science and Technology Research Center, Inc., 227-8502, Yokohama, Japan aut NATIONALLICENCE 700 1- China Hideyasu Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut NATIONALLICENCE 700 1- Saito Shigeki Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut NATIONALLICENCE 700 1- Kawabata Hiroshi Mitsubishi Chemical Group Science and Technology Research Center, Inc., 227-8502, Yokohama, Japan aut NATIONALLICENCE 700 1- Mihara Hisaaki Department of Biotechnology, College of Life Sciences, Ritsumeikan University, 525-8577, Kusatsu, Shiga, Japan aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/12(2015-06-01), 5045-5054 0175-7598 99:12<5045 2015 99 253