Characterization of a Glutamate Decarboxylase (GAD) from Enterococcus avium M5 Isolated from Jeotgal, a Korean Fermented Seafood
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To develop starters for the production of functional foods or materials, lactic acid bacteria producing γ-aminobutyric acid (GABA) were screened from jeotgals, Korean fermented seafoods. One isolate producing a high amount of GABA from monosodium L-glutamate (MSG) was identified as Enterococcus avium by 16S rRNA gene sequencing. E. avium M5 produced 18.47 ± 1.26 mg/ml GABA when incubated for 48 h at 37oC in MRS broth with MSG (3% (w/v)). A gadB gene encoding glutamate decarboxylase (GAD) was cloned and overexpressed in E. coli BL21 (DE3) using the pET26b (+) expression vector. Recombinant GAD was purified through a Ni-NTA column and the size was estimated to be 53 kDa by SDSPAGE. Maximum GAD activity was observed at pH 4.5 and 55oC and the activity was dependent on pyridoxal 5’-phosphate. The Km and Vmax values of GAD were 3.26 ± 0.21 mM and 0.0120 ± 0.0001 mM/min, respectively, when MSG was used as a substrate. Enterococcus avium M5 secretes a lot of GABA when grown on MRS with MSG, and the strain is useful for the production of fermented foods containing a high amount of GABA.
Abstract
To develop starters for the production of functional foods or materials, lactic acid bacteria producing γ-aminobutyric acid (GABA) were screened from jeotgals, Korean fermented seafoods. One isolate producing a high amount of GABA from monosodium L-glutamate (MSG) was identified as Enterococcus avium by 16S rRNA gene sequencing. E. avium M5 produced 18.47 ± 1.26 mg/ml GABA when incubated for 48 h at 37oC in MRS broth with MSG (3% (w/v)). A gadB gene encoding glutamate decarboxylase (GAD) was cloned and overexpressed in E. coli BL21 (DE3) using the pET26b (+) expression vector. Recombinant GAD was purified through a Ni-NTA column and the size was estimated to be 53 kDa by SDSPAGE. Maximum GAD activity was observed at pH 4.5 and 55oC and the activity was dependent on pyridoxal 5’-phosphate. The Km and Vmax values of GAD were 3.26 ± 0.21 mM and 0.0120 ± 0.0001 mM/min, respectively, when MSG was used as a substrate. Enterococcus avium M5 secretes a lot of GABA when grown on MRS with MSG, and the strain is useful for the production of fermented foods containing a high amount of GABA.
참고문헌 (27)
- Manyam BV, "Isoniazid-induced elevation of cerebrospinal fluid (CSF)GABA levels and effects on chorea in Huntington’s disease", Ann. Neurol, 10, pp.35~37 (1981)
- Inoue K, "Blood-pressure-lowering effect of a novel fermented milk containing γ-aminobutyric acid (GABA) in mild hypertensives", Eur. J. Clin. Nutr, 57, pp.490~495 (2003)
- Jakobs C, "Inherited disorders of GABA metabolism", J. Inherit. Metab. Dis, 16, pp.704~715 (1993)
- Wong CG, "GABA, γ-aminobutyric acid, and neurological disease", Ann. Neurol, 54, pp.S3~S12 (2003)
- Park KB, "Cloning, sequencing and expression of a novel glutamate decarboxylase gene from a newly isolated lactic acid bacterium, Lactobacillus brevis OPK-3", Bioresour. Technol, 98, pp.312~319 (2007)
- Li H, "Lactic acid bacteria cell factories for gamma-aminobutyric acid", Amino acids, 39, pp.1107~1116 (2010)
- Masuda K, "Isolation of marine yeasts collected from the Pacific-ocean showing a high production of γ-aminobutyric acid", Biosci. Biotechnol. Biochem, 72, pp.3265~3272 (2008)
- Leory F, "Lactic acid bacteria as functional starter cultures for the food fermentation industry", Trends Food Sci. Technol, 15, pp.67~78 (2004)
- Ueno H., "Enzymatic and structural aspects on glutamate decarboxylase", J. Mol. Catal. B Enzym, 10, pp.67~79 (2000)
- Park JY, "Characterization of a glutamate decarboxylase (GAD) gene from Lactobacillus zymae", Biotechnol. Lett, 36, pp.1791~1799 (2014)
- Tamura K, "MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods", Mol. Biol. Evol, 28, pp.2731~2739 (2011)
- Saitou N, "The neighbor-joining method: a new method for reconstructing phylogenetic trees", Mol. Biol. Evol, 4, pp.406~425 (1987)
- Tamura T, "Establishment of an efficient fermentation system of gamma-aminobutyric acid by a lactic acid bacterium, Enterococcus avium G-15, isolated from carrot leaves", Biol. Pharm. Bull, 33, pp.1673~1679 (2010)
- Hiraga K, "Glutamate decarboxylase from Lactobacillus brevis: activation by ammonium sulfate", Biosci. Biotechnol. Biochem, 72, pp.1299~1306 (2008)
- Nomura M, "Production of γ-aminobutyric acid by cheese starters during cheese ripening", J. Dairy Sci, 81, pp.1486~1491 (1998)
- Yokoyama S, "Production of γ-aminobutyric acid from alcohol distillery lees by Lactobacillus brevis IFO-12005", J. Biosci. Bioeng, 93, pp.95~97 (2002)
- Siragusa S, "Synthesis of γ-aminobutyric acid by lactic acid bacteria isolated from a variety of Italian cheeses", Appl. Environ. Microbiol, 73, pp.7283~7290 (2007)
- 사현덕, "Characterization of Glutamate Decarboxylase (GAD) from Lactobacillus sakei A156 Isolated from Jeot-gal", Journal of Microbiology and Biotechnology, 25(5), pp.696~703 (2015), 한국미생물·생명공학회[DOI]
- 국무창, "Enhanced Production of γ-Aminobutyric Acid Using Rice Bran Extracts by Lactobacillus sakei B2-16", Journal of Microbiology and Biotechnology, 20(4), pp.763~766 (2010), 한국미생물·생명공학회
- 임희선, "Optimization of γ-Aminobutyric Acid Production by Enterococcus faecium JK29 Isolated from a Traditional Fermented Foods", 한국미생물·생명공학회지, 44(1), pp.26~33 (2016), 한국미생물·생명공학회[DOI]
- Murzin AG., "Structural classification of proteins: new super families", Curr. Opin. Struct. Biol, 6, pp.386~394 (1996)
- Fonda ML., "L-Glutamate decarboxylase from bacteria", Methods Enzymol, 113, pp.11~16 (1985)
- Fan E, "Cloning, sequencing and expression of a glutamate decarboxylase gene from the GABA-producing strain Lactobacillus brevis CGMCC1306", Ann. Microbiol, 62, pp.689~698 (2012)
- Komatsuzaki N, "Characterization of glutamate decarboxylase from a high γ-aminobutyric acid (GABA)-producer, Lactobacillus paracasei", Biosci. Biotechnol. Biochem, 72, pp.278~285 (2008)
- Lin Q, "Cloning and expression of glutamate decarboxylase gene from Streptococcus thermophiles Y2", J. Gen. Appl. Microbiol, 55, pp.305~310 (2009)
- Yang H, "Accumulation of γ-aminobutyric acid by Enterococcus avium 9184 in scallop solution in a two-stage fermentation strategy", Microb. Biotechnol, 9, pp.478~485 (2015)
- Nomura M, "Lactococcus lactis contains only one glutamate decarboxylase gene", Microbiology, 145, pp.1375~1380 (1999)