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Characterization and large-scale production of recombinant Streptoverticillium platensis transglutaminase
- Lin, Shie-Jea, Hsieh, Yi-Fang, Lai, Li-An, Chao, Mei-Li, Chu, Wen-Shen
- Journal of industrial microbiology & biotechnology 2008 v.35 no.9 pp. 981-990
- EDTA (chelating agent), Streptomyces lividans, Streptomyces platensis, aluminum, ammonium sulfate, barium, calcium, cobalt, copper, enzyme activity, fermentation, fermenters, indole acetic acid, iron, lead, lithium, magnesium, manganese, mercury, molecular weight, pH, polyacrylamide gel electrophoresis, potassium, protein-glutamine gamma-glutamyltransferase, sodium, temperature, zinc
- Recombinant Streptomyces platensis transglutaminase (MtgA) produced by the Streptomyces lividans transformant 25-2 was purified by ammonium sulfate fractionation, followed by CM-Sepharose CL-6B fast flow, and blue-Sepharose fast flow chromatography. The purification factor was ~33.2-fold, and the yield was 65%. The molecular weight of the purified recombinant MtgA was 40.0 KDa as estimated by SDS-PAGE. The optimal pH and the temperature for the enzyme activity were 6.0 and 55 °C, respectively, and the enzyme was stable at pH 5.0-6.0 and at temperature 45-55 °C. Enzyme activity was not affected by Ca²⁺, Li⁺, Mn²⁺, Na⁺, Fe³⁺, K⁺, Mg²⁺, Al³⁺, Ba²⁺, Co²⁺, EDTA, or IAA but was inhibited by Fe²⁺, Pb²⁺, Zn²⁺, Cu²⁺, Hg²⁺, PCMB, NEM, and PMSF. Optimization of the fermentation medium resulted in a twofold increase of recombinant MtgA activity in both flasks (5.78 U/ml) and 5-l fermenters (5.39 U/ml). Large-scale productions of the recombinant MtgA in a 30-l air-lift fermenter and a 250-l stirred-tank fermenter were fulfilled with maximal activities of 5.36 and 2.54 U/ml, respectively.