FOOD SCIENCE ›› 2013, Vol. 34 ›› Issue (9): 90-94.doi: 10.7506/spkx1002-6630-201309020

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Intein-mediated Activation of Transglutaminase from Streptomyces hygroscopicus

DU Kun,ZHOU Li,DU Guo-cheng,CHEN Jian,ZHOU Zhe-min   

  1. 1. Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, China; 2. State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China
  • Received:2013-02-25 Revised:2013-04-14 Online:2013-05-15 Published:2013-05-07

Abstract: Microbial transglutaminase (MTG) from Streptomyces is secreted as an inactive form of pro-MTG, which can be activated by protease. However, protease has the potential for the hydrolysis of MTG as well as its substrates and can improve product separation costs. In this study, a pH-dependent mini-intein was inserted between the pro-region and MTG to mediate the cleavage of both parts by controlling pH. The recombinant protein (pro-Intein-MTG) was successfully overexpressed in Escherichia coli. The time required for converting recombinant protein to MTG was determined to be 24 h at 25 ℃ and pH 7.0 in vitro, resulting in an activity of 0.23 U/mL (cell concentration was diluted to OD600nm at 1.0). The specific activity and Km of the recombinant MTG was determined to be 14.3 U/mg and 69.4 mmol/L, respectively, which was consistent with that of wild type MTG from Streptomyces hygroscopicus. In addition, circular dichroism analysis showed that the recombinant and wild type MTG had similar secondary structure. These results indicated that the insertion of intein between the pro-region and MTG did not significantly affect the structure and function of MTG. This method is simple, economic and effective in the activation of pro-enzyme and has a great potential for industrial application.

Key words: protein splicing, intein, pro-enzyme activation, transglutaminase, Streptomyces hygroscopicus

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