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KMID : 0545120170270071216
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Journal of Microbiology and Biotechnology
2017 Volume.27 No. 7 p.1216 ~ p.1222
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Characterization of a Glutamate Decarboxylase (GAD) from Enterococcus avium M5 Isolated from Jeotgal, a Korean Fermented Seafood
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Lee Kang-Wook
Shim Jae-Min
Yao Zhuang
Kim Jeong-A
Kim Hyun-Jin
Kim Jeong-Hwan
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Abstract
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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.
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KEYWORD
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Gamma-aminobutyric acid, glutamate decarboxylase, gad gene cloning, Enterococcus avium, jeotgal
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