title: Application of repeated aspartate tags to improving extracellular production of Escherichia coli L-asparaginase isozyme II.
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.
authors: Department of Agricultural Biotechnology and Center for Food and Bioconvergence, Seoul National University, Seoul 151-921, South Korea.
authors: Sun-Ki
authors: SK
authors: Kim
authors: Department of Agricultural Biotechnology and Center for Food and Bioconvergence, Seoul National University, Seoul 151-921, South Korea.
authors: Won-Ki
authors: WK
authors: Min
authors: Department of Bio and Fermentation Convergence Technology, Kookmin University, Seoul 136-702, South Korea.
authors: Yong-Cheol
authors: YC
authors: Park
authors: Department of Agricultural Biotechnology and Center for Food and Bioconvergence, Seoul National University, Seoul 151-921, South Korea. Electronic address: jhseo94@snu.ac.kr.
authors: Jin-Ho
authors: JH
authors: Seo
mesh_terms: D001215:Asparaginase / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D001224:Aspartic Acid / Q000235:genetics; D001709:Biotechnology; D003470:Culture Media; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D005285:Fermentation; D017353:Gene Deletion; D005798:Genes, Bacterial; D007527:Isoenzymes / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D016324:Sequence Tagged Sites; D020079:Terminal Repeat Sequences
chemical_list: D003470:Culture Media; D029968:Escherichia coli Proteins; D007527:Isoenzymes; D011993:Recombinant Fusion Proteins; D001224:Aspartic Acid; D001215:Asparaginase
keywords: Escherichia coli; Extracellular production; General secretion pathway; PelB signal sequence; Repeated aspartate tag; l-Asparaginase isozyme II