An efficient protocol to enhance the extracellular production of recombinant protein from Escherichia coli by the synergistic effects of sucrose, glycine, and Triton X-100.

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  • Additional Information
    • Source:
      Publisher: Academic Press Country of Publication: United States NLM ID: 9101496 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1096-0279 (Electronic) Linking ISSN: 10465928 NLM ISO Abbreviation: Protein Expr Purif Subsets: MEDLINE
    • Publication Information:
      Publication: Orlando, FL : Academic Press
      Original Publication: San Diego : Academic Press, c1990-
    • Subject Terms:
      Gene Expression*; Alkaline Phosphatase/*biosynthesis ; Escherichia coli/*metabolism ; Glycine/*pharmacology ; Octoxynol/*pharmacology ; Periplasm/*metabolism ; Sucrose/*pharmacology; Alkaline Phosphatase/genetics ; Escherichia coli/genetics ; Periplasm/genetics ; Recombinant Fusion Proteins/biosynthesis ; Recombinant Fusion Proteins/genetics
    • Abstract:
      Targeting recombinant proteins at highly extracellular production in the culture medium of Escherichia coli presents a significant advantage over cytoplasmic or periplasmic expression. In this work, a recombinant protein between ZZ protein and alkaline phosphatase (rZZ-AP) was constructed. Because rZZ-AP has the IgG-binding capacity and enzymatic activity, it can serve as an immunoreagent in immunoassays. However, only a very small portion of rZZ-AP is generally secreted into the aqueous medium under conventional cultivation procedure. Hence, we emphasized on the optimization of the culture procedures and attempted to dramatically enhance the yield of extracellular rZZ-AP from E. coli HB101 host cells by adding sucrose, glycine, and Triton X-100 in the culture medium. Results showed that the extracellular production of rZZ-AP in the culture medium containing 5% sucrose, 1% glycine, and 1% Triton X-100 was 18.6 mg/l, which was 18.6-fold higher than that without the three chemicals. And the β-galactosidase activity test showed that the increased extracellular rZZ-AP was not due to cell lysis. Further analysis suggested a significant interaction effect among the three chemicals for the enhancement of extracellular production. Ultrastructural analysis indicated that the enhancement may be due to the influence of sucrose, glycine, and Triton X-100 on the periplasmic osmolality, permeability, or integrity of the cell wall, respectively. This proposed approach presents a simple strategy to enhance the extracellular secretion of recombinant proteins in the E. coli system at the process of cell cultivation.
      (Copyright © 2016 Elsevier Inc. All rights reserved.)
    • Contributed Indexing:
      Keywords: Escherichia coli; Extracellular secretion; Glycine; Sucrose; Synergistic effect; Triton X-100; ZZ protein-alkaline phosphatase fusion protein
    • Accession Number:
      0 (Recombinant Fusion Proteins)
      57-50-1 (Sucrose)
      9002-93-1 (Octoxynol)
      EC 3.1.3.1 (Alkaline Phosphatase)
      TE7660XO1C (Glycine)
    • Publication Date:
      Date Created: 20160519 Date Completed: 20170630 Latest Revision: 20220408
    • Publication Date:
      20240829
    • Accession Number:
      10.1016/j.pep.2016.05.007
    • Accession Number:
      27189822