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http://dx.doi.org/10.4014/jmb.1910.10044

Production of ʟ-Theanine Using Escherichia coli Whole-Cell Overexpressing γ-Glutamylmethylamide Synthetase with Baker's Yeast  

Yang, Soo-Yeon (Department of Biological Engineering, College of Engineering, Konkuk University)
Han, Yeong-Hoon (Department of Biological Engineering, College of Engineering, Konkuk University)
Park, Ye-Lim (Department of Biological Engineering, College of Engineering, Konkuk University)
Park, Jun-Young (Department of Biological Engineering, College of Engineering, Konkuk University)
No, So-young (Department of Biological Engineering, College of Engineering, Konkuk University)
Jeong, Daham (Department of Bioscience and Biotechnology, Konkuk University)
Park, Saerom (Department of Biological Engineering, College of Engineering, Konkuk University)
Park, Hyung Yeon (Department of Biological Engineering, College of Engineering, Konkuk University)
Kim, Wooseong (College of Pharmacy and Graduate School of Pharmaceutical Sciences, Ewha Womans University Seoul)
Seo, Seung-Oh (Department of Food Science and Nutrition, The Catholic University of Korea)
Yang, Yung-Hun (Department of Biological Engineering, College of Engineering, Konkuk University)
Publication Information
Journal of Microbiology and Biotechnology / v.30, no.5, 2020 , pp. 785-792 More about this Journal
Abstract
ʟ-Theanine, found in green tea leaves has been shown to positively affect immunity and relaxation in humans. There have been many attempts to produce ʟ-theanine through enzymatic synthesis to overcome the limitations of traditional methods. Among the many genes coding for enzymes in the ʟ-theanine biosynthesis, glutamylmethylamide synthetase (GMAS) exhibits the greatest possibility of producing large amounts of production. Thus, GMAS from Methylovorus mays No. 9 was overexpressed in several strains including vectors with different copy numbers. BW25113(DE3) cells containing the pET24ma::gmas was selected for strains. The optimal temperature, pH, and metal ion concentration were 50℃, 7, and 5 mM MnCl2, respectively. Additionally, ATP was found to be an important factor for producing high concentration of ʟ-theanine so several strains were tested during the reaction for ATP regeneration. Baker's yeast was found to decrease the demand for ATP most effectively. Addition of potassium phosphate source was demonstrated by producing 4-fold higher ʟ-theanine. To enhance the conversion yield, GMAS was additionally overexpressed in the system. A maximum of 198 mM ʟ-theanine was produced with 16.5 mmol/l/h productivity. The whole-cell reaction involving GMAS has greatest potential for scale-up production of ʟ-theanine.
Keywords
${\tiny\text{L}}$-theanine; whole-cell biocatalyst; baker's yeast; ATP regeneration;
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