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Production of γ-Aminobutyric Acid Using Immobilized Glutamate Decarboxylase from Lactobacillus plantarum

Lactobacillus plantarum 유래 글루탐산 탈탄산효소의 고정화를 이용한 γ-aminobutyric acid의 생산

  • 이상재 (신라대학교 식품공학전공) ;
  • 이한승 (신라대학교 식품공학전공) ;
  • 이동우 (경북대학교 응용생명과학부)
  • Received : 2015.09.03
  • Accepted : 2015.09.06
  • Published : 2015.09.28

Abstract

The glutamate decarboxylase gene (gadB) from Lactobacillus plantarum WCFS1 was cloned and expressed as an N-terminal hexa-histidine-tagged fusion protein in Escherichia coli BL21 (DE3) as the host strain. Purified glutamate decarboxylase (GAD) was immobilized onto porous silica beads by covalent coupling. The pH dependence of activity and stability of the immobilized GAD was significantly altered, when compared to those of the free enzyme. Immobilized GAD was stable in the range of pH 3.5 to 6.0. The resulting packed-bed reactor produced 41.7 g of γ-aminobutyric acid/l·h at 45℃.

효율적인 γ-aminobutyric acid (GABA)의 생산을 위해 Lactobacillus plantarum WCFS1로부터 글루탐산 탈탄산효소(glutamate decarboxylase, GAD)를 대장균에 발현, 정제 후 silica beads에 covalent coupling 방법을 이용하여 고정화하였다. 고정화된 효소의 특성을 고정화하지 않은 효소와 비교한 결과, 모든 pH의 범위(pH 3.5–6.0)에서 80% 이상의 활성을 나타내었으며 pH 안정성과 열 안정성 모두 증대되었다. 이 고정화 효소를 packed-bed reactor에 충진하여 GABA의 생산성을 확인한 결과 1리터당 1시간에 최대 41.7 g의 GABA 생산이 가능한 것으로 확인되었다.

Keywords

References

  1. Albayrak N, Yang ST. 2002. Immobilization of beta-galactosidase on fibrous matrix by polyethyleneimine for production of galacto-oligosaccharides from lactose. Biotechnol. Prog. 18: 240−251. https://doi.org/10.1021/bp010167b
  2. Arica MY, Hasirci V, Alaeddinoglu NG. 1995. Covalent immobilization of alpha-amylase onto pHEMA microspheres: preparation and application to fixed bed reactor. Biomaterials. 16: 761−768. https://doi.org/10.1016/0142-9612(95)99638-3
  3. Barrett E, Ross RP, O’Toole PW, Fitzgerald GF, Stanton C. 2012. gamma-Aminobutyric acid production by culturable bacteria from the human intestine. J. Appl. Microbiol. 113: 411−417. https://doi.org/10.1111/j.1365-2672.2012.05344.x
  4. Blankenhorn D, Phillips J, Slonczewski JL. 1999. Acid- and base-induced proteins during aerobic and anaerobic growth of Escherichia coli revealed by two-dimensional gel electrophoresis. J. Bacteriol. 181: 2209−2216.
  5. Busto MD. 1998. An experiment illustrating the effect of immobilisation on enzyme properties. Biochem. Edu. 26: 304−308. https://doi.org/10.1016/S0307-4412(98)00168-X
  6. De Biase D, Tramonti A, Bossa F, Visca P. 1999. The response to stationary-phase stress conditions in Escherichia coli: role and regulation of the glutamic acid decarboxylase system. Mol. Microbiol. 32: 1198−1211. https://doi.org/10.1046/j.1365-2958.1999.01430.x
  7. Di Cagno R, Mazzacane F, Rizzello CG, De Angelis M, Giuliani G, Meloni M, et al. 2010. Synthesis of gamma-aminobutyric acid (GABA) by Lactobacillus plantarum DSM19463: functional grape must beverage and dermatological applications. Appl. Microbiol. Biotechnol. 86: 731−741. https://doi.org/10.1007/s00253-009-2370-4
  8. Foster AC, Kemp JA. 2006. Glutamate- and GABA-based CNS therapeutics. Curr. Opin. Pharmacol. 6: 7−17. https://doi.org/10.1016/j.coph.2005.11.005
  9. Jeng KC, Chen CS, Fang YP, Hou RC, Chen YS. 2007. Effect of microbial fermentation on content of statin, GABA, and polyphenols in Pu-Erh tea. J. Agric. Food Chem. 55: 8787−8792. https://doi.org/10.1021/jf071629p
  10. Kakee A, Takanaga H, Terasaki T, Naito M, Tsuruo T, Sugiyama Y. 2001. Efflux of a suppressive neurotransmitter, GABA, across the blood-brain barrier. J. Neurochem. 79: 110−118.
  11. Lee S, Ahn J, Kim YG, Jung JK, Lee H, Lee EG. 2013. Gamma-aminobutyric Acid production using immobilized glutamate decarboxylase followed by downstream processing with cation exchange chromatography. Int. J. Mol. Sci. 14: 1728−1739. https://doi.org/10.3390/ijms14011728
  12. Lin Q. 2013. Submerged fermentation of Lactobacillus rhamnosus YS9 for gamma-aminobutyric acid (GABA) production. Braz. J. Microbiol. 44: 183−187. https://doi.org/10.1590/S1517-83822013000100028
  13. Meldrum BS, Rogawski MA. 2007. Molecular targets for antiepileptic drug development. Neurotherapeutics. 4: 18−61. https://doi.org/10.1016/j.nurt.2006.11.010
  14. Nomura M, Nakajima I, Fujita Y, Kobayashi M, Kimoto H, Suzuki I, Aso H. 1999. Lactococcus lactis contains only one glutamate decarboxylase gene. Microbiology. 145: 1375−1380. https://doi.org/10.1099/13500872-145-6-1375
  15. Ortega N, Busto MD, Perez-Mateos M. 1998. Stabilisation of beta-glucosidase entrapped in alginate and polyacrylamide gels towards thermal and proteolytic deactivation. J. Chem. Technol. Biotechnol. 73: 7−12. https://doi.org/10.1002/(SICI)1097-4660(199809)73:1<7::AID-JCTB921>3.0.CO;2-#
  16. Rehm H-Jr, Reed G, Kennedy JF. 1987. Biotechnology. Vol. 7a, Enzyme Technology. VCH, Weinheim.
  17. Rizzello CG, Cassone A, Di Cagno R, Gobbetti M. 2008. Synthesis of angiotensin I-converting enzyme (ACE)-inhibitory peptides and gamma-aminobutyric acid (GABA) during sourdough fermentation by selected lactic acid bacteria. J. Agric. Food Chem. 56: 6936−6943. https://doi.org/10.1021/jf800512u
  18. Shin SM, Kim H, Joo Y, Lee SJ, Lee YJ, Lee SJ, et al. 2014. Characterization of glutamate decarboxylase from Lactobacillus plantarum and its C-terminal function for the pH dependence of activity. J. Agric. Food Chem. 62: 12186−12193. https://doi.org/10.1021/jf504656h
  19. Weetall HH. 1976. Covalent coupling methods for inorganic support materials. Methods Enzymol. 44: 134−148. https://doi.org/10.1016/S0076-6879(76)44012-0
  20. Yao W, Wu X, Zhu J, Sun B, Miller C. 2013. In vitro enzymatic conversion of gamma-aminobutyric acid immobilization of glutamate decarboxylase with bacterial cellulose membrane (BCM) and non-linear model establishment. Enzyme Microb. Technol. 52: 258−264. https://doi.org/10.1016/j.enzmictec.2013.01.008

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