Application of repeated aspartate tags to improving extracellular production of Escherichia coli l-asparaginase isozyme II |
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| Institution: | 1. Department of Agricultural Biotechnology and Center for Food and Bioconvergence, Seoul National University, Seoul 151-921, South Korea;2. Department of Bio and Fermentation Convergence Technology, Kookmin University, Seoul 136-702, South Korea;1. State Key Laboratory of Biotherapy/Collaborative Innovation Center for Biotherapy, West China Medical School, West China Hospital, Sichuan University, Chengdu, PR China;2. The Gastroenterology Tumor and Microenvironment Laboratory, Department of Gastroenterology, The First Affiliated Hospital of Chengdu Medical College, Chengdu Medical College, Chengdu, PR China;3. Chengdu Institute of Biological Products Co., Ltd., PR China;1. Departamento de Tecnologia Bioquímico-Farmacêutica, Faculdade de Ciências Farmacêuticas, Universidade de São Paulo, São Paulo, Brazil;2. Department of Chemical Engineering, Faculty of Engineering and Sciences, University of La Frontera, Temuco, Chile;3. Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo, Brazil |
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| Abstract: | Asparaginase isozyme II from Escherichia coli is a popular enzyme that has been used as a therapeutic agent against acute lymphoblastic leukemia. Here, fusion tag systems consisting of the pelB signal sequence and various lengths of repeated aspartate tags were devised to highly express and to release active asparaginase isozyme II extracellularly in E. coli. Among several constructs, recombinant asparaginase isozyme II fused with the pelB signal sequence and five aspartate tag was secreted efficiently into culture medium at 34.6 U/mg cell of specific activity. By batch fermentation, recombinant E. coli produced 40.8 U/ml asparaginase isozyme II in the medium. In addition, deletion of the gspDE gene reduced extracellular production of asparaginase isozyme II, indicating that secretion of recombinant asparaginase isozyme II was partially ascribed to the recognition by the general secretion machinery. This tag system composed of the pelB signal peptide, and repeated aspartates can be applied to extracellular production of other recombinant proteins. |
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| Keywords: | Extracellular production Repeated aspartate tag PelB signal sequence General secretion pathway |
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