Efficient secretory expression of recombinant proteins in Escherichia coli with a novel actinomycete signal peptide.

Item request has been placed! ×
Item request cannot be made. ×
loading   Processing Request
Read More Add to Saved list
  • 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:
      Bacterial Proteins*/biosynthesis ; Bacterial Proteins*/genetics ; Bacterial Secretion Systems*/genetics ; Bacterial Secretion Systems*/metabolism ; Escherichia coli*/genetics ; Escherichia coli*/metabolism ; Protein Sorting Signals* ; alpha-Amylases*/biosynthesis ; alpha-Amylases*/genetics; Actinobacteria/*genetics ; Bacillus subtilis/*genetics; Bacillus subtilis/enzymology ; Recombinant Proteins/biosynthesis ; Recombinant Proteins/genetics
    • Abstract:
      In well-established heterologous hosts, such as Escherichia coli, recombinant proteins are usually intracellular and frequently found as inclusion bodies-especially proteins possessing high rare codon content. In this study, successful secretory expression of three hydrolases, in a constructed inducible or constitutive system, was achieved by fusion with a novel signal peptide (Kp-SP) from an actinomycete. The signal peptide efficiently enabled extracellular protein secretion and also contributed to the active expression of the intracellular recombinant proteins. The thermophilic α-amylase gene of Bacillus licheniformis was fused with Kp-SP. Both recombinants, carrying inducible and constitutive plasmids, showed remarkable increases in extracellular and intracellular amylolytic activity. Amylase activity was observed to be > 10-fold in recombinant cultures with the constitutive plasmid, pBSPPc, compared to that in recombinants lacking Kp-SP. Further, the signal peptide enabled efficient secretion of a thermophilic cellulase into the culture medium, as demonstrated by larger halo zones and increased enzymatic activities detected in both constructs from different plasmids. For heterologous proteins with a high proportion of rare codons, it is difficult to obtain high expression in E. coli owing to the codon bias. Here, the fusion of an archaeal homologue of the amylase encoding gene, FSA, with Kp-SP resulted in > 5-fold higher extracellular activity. The successful extracellular expression of the amylase indicated that the signal peptide also contributed significantly to its active expression and signified the potential value of this novel and versatile signal peptide in recombinant protein production.
      (Copyright © 2016 Elsevier Inc. All rights reserved.)
    • Contributed Indexing:
      Keywords: Actinomycete signal peptide; Extracellular expression; High rare codon content protein; Inducible and constitutive expression
    • Accession Number:
      0 (Bacterial Proteins)
      0 (Bacterial Secretion Systems)
      0 (Protein Sorting Signals)
      0 (Recombinant Proteins)
      EC 3.2.1.1 (alpha-Amylases)
    • Publication Date:
      Date Created: 20160925 Date Completed: 20170630 Latest Revision: 20201209
    • Publication Date:
      20231215
    • Accession Number:
      10.1016/j.pep.2016.09.011
    • Accession Number:
      27664436