Characterization of glutamate decarboxylase from a high gamma-aminobutyric acid (GABA)-producer, Lactobacillus paracasei.
Gamma-aminobutyric acid (GABA) has several physiological functions in humans. We have reported that Lactobacillus paracasei NFRI 7415 produces high levels of GABA. To gain insight into the higher GABA-producing ability of this strain, we analyzed glutamate decarboxylase (GAD), which catalyzes the decarboxylation of L-glutamate to GABA. The molecular weight of the purified GAD was estimated to be 57 kDa by SDS-PAGE and 110 kDa by gel filtration, suggesting that GAD forms the dimer under native conditions. GAD activity was optimal at pH 5.0 at 50 degrees C. The Km value for the catalysis of glutamate was 5.0 mM, and the maximum rate of catalysis was 7.5 micromol min(-1) mg(-1). The N-terminal amino acid sequence of GAD was determined, and the gene encoding GAD from genomic DNA was cloned. The findings suggest that the ability of Lb. paracasei to produce high levels of GABA results from two characteristics of GAD, viz., a low Km value and activity at low pH.
Komatsuzaki
Noriko
N
National Food Research Institute, Tsukuba, Ibaraki 305-8642, Japan.
Nakamura
Toshihide
T
Kimura
Toshinori
T
Shima
Jun
J
D000595:Amino Acid Sequence; D001483:Base Sequence; D002384:Catalysis; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D017931:DNA Primers; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D005798:Genes, Bacterial; D005968:Glutamate Decarboxylase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007778:Lactobacillus / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013696:Temperature; D005680:gamma-Aminobutyric Acid / Q000096:biosynthesis*
D017931:DNA Primers; D005680:gamma-Aminobutyric Acid; D005968:Glutamate Decarboxylase