Abstract
A putative gene (gadlbhye1) encoding glutamate decarboxylase (GAD) was cloned from Lactobacillus brevis HYE1 isolated from kimchi, a traditional Korean fermented vegetable. The amino acid sequences of GADLbHYE1 showed 48% homology with the GadA family and 99% identity with the GadB family from L. brevis. The cloned GADLbHYE1 was functionally expressed in Escherichia coli using inducible expression vectors. The expressed recombinant GADLbHYE1 was successfully purified by Ni–NTA affinity chromatography, and had a molecular mass of 54 kDa with optimal hydrolysis activity at 55 °C and pH 4.0. Its thermal stability was determined to be higher than that of other GADs from L. brevis, based on its melting temperature (75.18 °C). Kinetic parameters including Km and Vmax values for GADLbHYE1 were 4.99 mmol/L and 0.224 mmol/L/min, respectively. In addition, the production of gamma-aminobutyric acid in E. coli BL21 harboring gadlbhye1/pET28a was increased by adding pyridoxine as a cheaper coenzyme.
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Funding
This work was supported by Main Research Programs of the Korea Food Research Institute (KFRI) funded by the Ministry of Science, ICT & Future Planning [Grant Number E0170602-01]; Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT & Future Planning [Grant Number NRF-2014R1A1A1002980]; Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education [Grant Number NRF-2016R1D1A1B03931582].
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All authors conceived the experiments and discussed the results. HSL and HL performed the experiments and analyzed the data with D-HS, I-TC and M-JS. D-HS, Y-DN and M-JS contributed to the writing of the paper.
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Lim, H.S., Seo, DH., Cha, IT. et al. Expression and characterization of glutamate decarboxylase from Lactobacillus brevis HYE1 isolated from kimchi. World J Microbiol Biotechnol 34, 44 (2018). https://doi.org/10.1007/s11274-018-2427-6
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DOI: https://doi.org/10.1007/s11274-018-2427-6