title: Characterization of Glutamate Decarboxylase (GAD) from Lactobacillus sakei A156 Isolated from Jeot-gal.
abstract: A gamma-aminobutyric acid (GABA)-producing microorganism was isolated from jeot-gal (anchovy), a Korean fermented seafood. The isolate, A156, produced GABA profusely when incubated in MRS broth with monosodium glutamate (3% (w/v)) at 37°C for 48 h. A156 was identified as Lactobacillus sakei by 16S rRNA gene sequencing. The GABA conversion yield was 86% as determined by GABase enzyme assay. The gadB gene encoding glutamate decarboxylase (GAD) was cloned by PCR. gadC encoding a glutamate/GABA antiporter was located immediately upstream of gadB. The operon structure of gadCB was confirmed by RT-PCR. gadB was overexpressed in Escherichia coli BL21(DE3) and recombinant GAD was purified. The purified GAD was 54.4 kDa in size by SDS-PAGE. Maximum GAD activity was observed at pH 5.0 and 55°C and the activity was dependent on pyridoxal 5'-phosphate. The Km and Vmax of GAD were 0.045 mM and 0.011 mM/min, respectively, when glutamate was used as the substrate.
authors: Division of Applied Life Science (BK21 Plus), Graduate School, Gyeongsang National University, Jinju 660-701, Republic of Korea.
authors: Hyun Deok
authors: HD
authors: Sa
authors: Division of Applied Life Science (BK21 Plus), Graduate School, Gyeongsang National University, Jinju 660-701, Republic of Korea.
authors: Ji Yeong
authors: JY
authors: Park
authors: Division of Applied Life Science (BK21 Plus), Graduate School, Gyeongsang National University, Jinju 660-701, Republic of Korea.
authors: Seon-Ju
authors: SJ
authors: Jeong
authors: Institute of Agriculture and Life Science, Gyeongsang National University, Jinju 660-701, Republic of Korea.
authors: Kang Wook
authors: KW
authors: Lee
authors: Division of Applied Life Science (BK21 Plus), Graduate School, Gyeongsang National University, Jinju 660-701, Republic of Korea.|Institute of Agriculture and Life Science, Gyeongsang National University, Jinju 660-701, Republic of Korea.
authors: Jeong Hwan
authors: JH
authors: Kim
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D005285:Fermentation; D005968:Glutamate Decarboxylase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007778:Lactobacillus / Q000201:enzymology* / Q000235:genetics; D058977:Molecular Sequence Annotation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D056910:Republic of Korea; D017747:Seafood / Q000382:microbiology*; D016415:Sequence Alignment; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D011994:Recombinant Proteins; D005680:gamma-Aminobutyric Acid; D005968:Glutamate Decarboxylase