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Gene expression and characterization of thermostable glutamate decarboxylase from Pyrococcus furiosus

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Abstract

Gene encoding for a putative glutamate decarboxylase (GAD: EC 4.1.1.15) from the hyperthermophilic archaeon Pyrococcus furiosus was cloned and the biochemical characteristics of the resulting recombinant protein were examined. The gene (PF1159) from P. furiosus showed some identity with other group II decarboxylases from an archaea and bacteria. The GAD from P. furiosus (PfGAD) was expressed in Escherichia coli, and the recombinant protein has a molecular mass of 41 kDa, determined by SDS-PAGE. The optimum temperature and pH for GAD activity were 75°C and 6.0, respectively. The half-life of heat inactivation was approximately 60 min at 90°C. The GAD activity was found to be dependent on various salts, such as CaCl2, NaCl, KCl, and NaBr, with an optimum concentration of 400 mM, but not (NH4)2SO4. PfGAD demonstrated activity against various substrates, such as l-glutamate, l-aspartate, and l-tyrosine. The results of the kinetics experiment indicated that l-aspartate was a better substrate of PfGAD than l-glutamate and Ltyrosine.

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Authors and Affiliations

  1. Department of Biomaterial Control (BK21 program), Dong-Eui University, Busan, 614-714, Korea

    Eon-Seok Lee, Yeon-Hee Kim, Soo-Wan Nam, Byung-Woo Kim & Sung-Jong Jeon

  2. Department of Biotechnology & Bioengineering, Dong-Eui University, Busan, 614-714, Korea

    Yeon-Hee Kim, Soo-Wan Nam & Sung-Jong Jeon

  3. Department of Life Science and Biotechnology, Dong-Eui University, Busan, 614-714, Korea

    Byung-Woo Kim

  4. Blue-Bio Industry RIC, Dong-Eui University, Busan, 614-714, Korea

    Soo-Wan Nam, Byung-Woo Kim & Sung-Jong Jeon

  5. Division of Life Sciences, Korea Polar Research Institute, Incheon, 406-840, Korea

    Han-Woo Kim

  6. Department of Bioscience and Biotechnology, Konkuk University, Seoul, 143-701, Korea

    Dong-Eun Kim

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  1. Eon-Seok Lee
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Correspondence to Sung-Jong Jeon.

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Lee, ES., Kim, HW., Kim, DE. et al. Gene expression and characterization of thermostable glutamate decarboxylase from Pyrococcus furiosus . Biotechnol Bioproc E 18, 375–381 (2013). https://doi.org/10.1007/s12257-012-0581-5

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  • DOI: https://doi.org/10.1007/s12257-012-0581-5

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