Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
권호 표지
DOI QR코드

DOI QR Code

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)
  • Received : 2019.10.21
  • Accepted : 2020.02.21
  • Published : 2020.05.28
Download PDF
⟨ Previous Next ⟩

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

References

  1. Zheng G, Sayama K, Okubo T, Juneja LR, Oguni I. 2004. Anti-obesity effects of three major components of green tea, catechins, caffeine and theanine, in mice. In Vivo 18: 55-62.
  2. Turkozu D, Sanlier N. 2017. L-theanine, unique amino acid of tea, and its metabolism, health effects, and safety. Crit. Rev. Food Sci. Nutr. 57: 1681-1687. https://doi.org/10.1080/10408398.2015.1016141
  3. Saeed M, Naveed M, Arif M, Kakar MU, Manzoor R, Abd El-Hack ME, et al. 2017. Green tea (Camellia sinensis) and l-theanine: Medicinal values and beneficial applications in humans-A comprehensive review. Biomed. Pharmacother. 95: 1260-1275. https://doi.org/10.1016/j.biopha.2017.09.024
  4. Adhikary R, Mandal V. 2017. L-theanine: A potential multifaceted natural bioactive amide as health supplement. Asian Pac. J. Trop. Biomed. 7: 842-848. https://doi.org/10.1016/j.apjtb.2017.08.005
  5. Vuong QV, Stathopoulos CE, Golding JB, Nguyen MH, Roach PD. 2011. Optimum conditions for the water extraction of L-theanine from green tea. J. Sep. Sci. 34: 2468-2474. https://doi.org/10.1002/jssc.201100401
  6. Vuong QV, Bowyer MC, Roach PD. 2011. L-Theanine: properties, synthesis and isolation from tea. J. Sci. Food Agric. 91: 1931-1939. https://doi.org/10.1002/jsfa.4373
  7. Kawagishi H, Sugiyama K. 1992. Facile and large-scale synthesis of L-Theanine. Biosci. Biotechnol. Biochem. 56: 689. https://doi.org/10.1271/bbb.56.689
  8. Shuai Y, Zhang T, Jiang B, Mu W. 2010. Development of efficient enzymatic production of theanine by gamma-glutamyltranspeptidase from a newly isolated strain of Bacillus subtilis, SK11.004. J. Sci. Food Agric. 90: 2563-2567. https://doi.org/10.1002/jsfa.4120
  9. Tachiki T, Yamada T, Mizuno K, Ueda M, Shiode J, Fukami H. 1998. gamma-glutamyl transfer reactions by glutaminase from Pseudomonas nitroreducens IFO 12694 and their application for the syntheses of theanine and gamma-glutamylmethylamide. Biosci. Biotechnol. Biochem. 62: 1279-1283. https://doi.org/10.1271/bbb.62.1279
  10. Mu W, Zhang T, Jiang B. 2015. An overview of biological production of L-theanine. Biotechnol. Adv. 33: 335-342. https://doi.org/10.1016/j.biotechadv.2015.04.004
  11. Chen X, Su L, Wu D, Wu J. 2014. Application of recombinant Bacillus subtilis $\gamma$-glutamyltranspeptidase to the production of ltheanine. Process Biochem. 49: 1429-1439. https://doi.org/10.1016/j.procbio.2014.05.019
  12. Pu H, Wang Q, Zhu F, Cao X, Xin Y, Luo L, et al. 2013. Cloning, expression of glutaminase from Pseudomonas nitroreducens and application to theanine synthesis. Biocatal. Biotransform. 31: 1-7. https://doi.org/10.3109/10242422.2012.749462
  13. Sharma E, Joshi R, Gulati A. 2018. l-Theanine: An astounding sui generis integrant in tea. Food Chem. 242: 601-610. https://doi.org/10.1016/j.foodchem.2017.09.046
  14. Liu S, Li Y, Zhu J. 2016. Enzymatic production of l-theanine by $\gamma$-glutamylmethylamide synthetase coupling with an ATP regeneration system based on polyphosphate kinase. Process Biochem. 51: 1458-1463. https://doi.org/10.1016/j.procbio.2016.06.006
  15. Berke W, Schuz HJ, Wandrey C, Morr M, Denda G, Kula MR. 1988. Continuous regeneration of ATP in enzyme membrane reactor for enzymatic syntheses. Biotechnol. Bioeng. 32: 130-139. https://doi.org/10.1002/bit.260320203
  16. Endo T, Koizumi S. 2001. Microbial conversion with cofactor regeneration using genetically engineered bacteria. Adv. Synth. Catal. 343: 521-526. https://doi.org/10.1002/1615-4169(200108)343:6/7<521::AID-ADSC521>3.0.CO;2-5
  17. Moon Y-M, Yang SY, Choi TR, Jung H-R, Song H-S, hoon Han Y, et al. 2019. Enhanced production of cadaverine by the addition of hexadecyltrimethylammonium bromide to whole cell system with regeneration of pyridoxal-5'-phosphate and ATP. Enzyme Microb. Technol. 127: 58-64. https://doi.org/10.1016/j.enzmictec.2019.04.010
  18. Wei LL, Goux WJ. 1992. ATP cofactor regeneration via the glycolytic pathway. Bioorg. Chem. 20: 62-66. https://doi.org/10.1016/0045-2068(92)90026-y
  19. Sato M, Masuda Y, Kirimura K, Kino K. 2007. Thermostable ATP regeneration system using polyphosphate kinase from Thermosynechococcus elongatus BP-1 for D-amino acid dipeptide synthesis. J. Biosci. Bioeng. 103: 179-184. https://doi.org/10.1263/jbb.103.179
  20. Kameda A, Shiba T, Kawazoe Y, Satoh Y, Ihara Y, Munekata M, et al. 2001. A novel ATP regeneration system using polyphosphate-AMP phosphotransferase and polyphosphate kinase. J. Biosci. Bioeng. 91: 557-563. https://doi.org/10.1016/S1389-1723(01)80173-0
  21. Yan B, Ding Q, Ou L, Zou Z. 2014. Production of glucose-6-phosphate by glucokinase coupled with an ATP regeneration system. World J. Microbiol. Biotechnol. 30: 1123-1128. https://doi.org/10.1007/s11274-013-1534-7
  22. Resnick SM, Zehnder AJ. 2000. In vitro ATP regeneration from polyphosphate and AMP by polyphosphate: AMP phosphotransferase and adenylate kinase from Acinetobacter johnsonii 210A. Appl. Environ. Microbiol. 66: 2045-2051. https://doi.org/10.1128/AEM.66.5.2045-2051.2000
  23. Wakisaka S, Ohshima Y, Ogawa M, Tochikura T, Tachiki T. 1998. Characteristics and efficiency of glutamine production by coupling of a bacterial glutamine synthetase reaction with the alcoholic fermentation system of baker's yeast. Appl. Environ. Microbiol. 64: 2952-2957. https://doi.org/10.1128/aem.64.8.2952-2957.1998
  24. Horinouchi N, Sakai T, Kawano T, Matsumoto S, Sasaki M, Hibi M, et al. 2012. Construction of microbial platform for an energyrequiring bioprocess: practical 2'-deoxyribonucleoside production involving a C- C coupling reaction with high energy substrates. Microb. Cell Fact. 11: 82. https://doi.org/10.1186/1475-2859-11-82
  25. Matsuno R, Asada M, Nakanishi K, Kamikubo T. 1982. ATP Regeneration by Enzymes of Alcohol Fermentation and Kinases of Yeast and its Computer Simulation, pp. 351-352. Enzyme Engineering, Ed. Springer
  26. Horinouchi N, Ogawa J, Kawano T, Sakai T, Saito K, Matsumoto S, et al. 2006. Efficient production of 2-deoxyribose 5-phosphate from glucose and acetaldehyde by coupling of the alcoholic fermentation system of Baker's yeast and deoxyriboaldolase-expressing Escherichia coli. Biosci. Biotechnol. Biochem. 70: 1371-1378. https://doi.org/10.1271/bbb.50648
  27. Yamamoto S, Wakayama M, Tachiki T. 2005. Theanine production by coupled fermentation with energy transfer employing Pseudomonas taetrolens Y-30 glutamine synthetase and baker's yeast cells. Biosci. Biotechnol. Biochem. 69: 784-789. https://doi.org/10.1271/bbb.69.784
  28. Lin J-P, Tian J, You J-F, Jin Z-H, Xu Z-N, Cen P-L. 2004. An effective strategy for the co-production of S-adenosyl-L-methionine and glutathione by fed-batch fermentation. Biochem. Eng. J. 21: 19-25. https://doi.org/10.1016/j.bej.2004.04.013
  29. Bhatia SK, Bhatia RK, Yang Y-H. 2016. Biosynthesis of polyesters and polyamide building blocks using microbial fermentation and biotransformation. Rev. Environ. Sci. Bio/Technol. 15: 639-663. https://doi.org/10.1007/s11157-016-9415-9
  30. Yang S-Y, Choi T-R, Jung H-R, Park Y-L, Han Y-H, Song H-S, et al. 2020. Development of glutaric acid production consortium system with $\alpha$-ketoglutaric acid regeneration by glutamate oxidase in Escherichia coli. Enzyme Microb. Technol. 133: 109446. https://doi.org/10.1016/j.enzmictec.2019.109446
  31. Moon Y-M, Gurav R, Kim J, Hong Y-G, Bhatia SK, Jung H-R, et al. 2018. Whole-cell immobilization of engineered Escherichia coli JY001 with barium-alginate for itaconic acid production. Biotechnol. Bioprocess Eng. 23: 442-447. https://doi.org/10.1007/s12257-018-0170-3
  32. Bhatia SK, Shim Y-H, Jeon J-M, Brigham CJ, Kim Y-H, Kim H-J, et al. 2015. Starch based polyhydroxybutyrate production in engineered Escherichia coli. Bioprocess Biosys. Eng. 38: 1479-1484. https://doi.org/10.1007/s00449-015-1390-y
  33. Bhatia SK, Kim S-H, Yoon J-J, Yang Y-H. 2017. Current status and strategies for second generation biofuel production using microbial systems. Energy Convers. Manage 148: 1142-1156. https://doi.org/10.1016/j.enconman.2017.06.073
  34. Alaiz M, Navarro JL, Giron J, Vioque E. 1992. Amino acid analysis by high-performance liquid chromatography after derivatization with diethyl ethoxymethylenemalonate. J. Chromatogr. 591: 181-186. https://doi.org/10.1016/0021-9673(92)80236-N
  35. Kim J, Seo H-M, Bhatia SK, Song H-S, Kim J-H, Jeon J-M, et al. 2017. Production of itaconate by whole-cell bioconversion of citrate mediated by expression of multiple cis-aconitate decarboxylase (cadA) genes in Escherichia coli. Sci. Rep. 7: 39768. https://doi.org/10.1038/srep39768
  36. Griffiths MW, Muir DD. 1978. Properties of a thermostable $\beta$-galactosidase from a thermophilic Bacillus: Comparison of the enzyme activity of whole cells, purified enzyme and immobilised whole cells. J. Sci. Food Agric. 29: 753-761. https://doi.org/10.1002/jsfa.2740290904
  37. Buchachenko AL, Kuznetsov DA. 2008. Magnetic field affects enzymatic ATP synthesis. J. Am. Chem. Soc. 130: 12868-12869. https://doi.org/10.1021/ja804819k
  38. Moon Y-M, Yang SY, Choi TR, Jung H-R, Song H-S, hoon Han Y, et al. 2019. Enhanced production of cadaverine by the addition of hexadecyltrimethylammonium bromide to whole cell system with regeneration of pyridoxal-5'-phosphate and ATP. Enzyme Microb. Technol. 127: 58-64. https://doi.org/10.1016/j.enzmictec.2019.04.010
  39. Taylor RG, Walker DC, McInnes R. 1993. E. coli host strains significantly affect the quality of small scale plasmid DNA preparations used for sequencing. Nucleic Acids Res. 21: 1677. https://doi.org/10.1093/nar/21.7.1677
  40. Lobstein J, Emrich CA, Jeans C, Faulkner M, Riggs P, Berkmen M. 2012. SHuffle, a novel Escherichia coli protein expression strain capable of correctly folding disulfide bonded proteins in its cytoplasm. Microb. Cell Fact. 11: 753. https://doi.org/10.1186/1475-2859-11-56
  41. Lee SG, Lee JO, Yi JK, Kim BG. 2002. Production of cytidine 5'-monophosphate N-acetylneuraminic acid using recombinant Escherichia coli as a biocatalyst. Biotechnol. Bioeng. 80: 516-524. https://doi.org/10.1002/bit.10398
  42. Jakoby M, Ngouoto-Nkili C-E, Burkovski A. 1999. Construction and application of new Corynebacterium glutamicum vectors. Biotechnol. Tech. 13: 437-441. https://doi.org/10.1023/A:1008968419217

Cited by

  1. Overview on Multienzymatic Cascades for the Production of Non-canonical α-Amino Acids vol.8, 2020, https://doi.org/10.3389/fbioe.2020.00887
  2. New advances in genetic engineering for L-theanine biosynthesis vol.114, 2021, https://doi.org/10.1016/j.tifs.2021.06.006
  3. Metabolic Engineering of Pseudomonas putida for Fermentative Production of L-Theanine vol.69, pp.34, 2020, https://doi.org/10.1021/acs.jafc.1c03240