Production of antimicrobial peptide DLP4 in Escherichia coli using an ELP-Intein system
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    Abstract:

    DLP4 (defensin-like peptide 4) is a novel insect defensin, which has strong antibacterial activity against Gram-positive bacteria and is not susceptible to develop drug resistance. In this study, an elastin-like polypeptide (ELP) and an intein fusion system were used for production and purification of DLP4, which combined the characteristics of the phase transition of ELP and the C-cleavage of the intein. A recombinant expression plasmid pET-ELP-I-DLP4 was constructed and transformed into Escherichia coli. Subsequently, DLP4 was purified by simple centrifugation, alternation of pH and temperature. However, the C-cleavage of the intein occurred unexpectedly during the process of expression and purification. To solve this problem, the intein was split into N-intein (I0N) and C-intein (I0C), and fused with ELP or DLP4 to construct two recombinant expression plasmids pET-ELP-I0N and pET-ELP-I0C-DLP4, respectively. These two plasmids were transformed into E. coli separately. The mixture of the two cultures of E. coli strains restored the C-cleavage activity of the intein. This operation yielded DLP4 of about 1.49 mg/L. Antibacterial test confirmed that the purified DLP4 exhibited expected activity. Thus, this approach can be used as an effective way for DLP4 expression and purification in the prokaryotic system.

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姜宇,李秀,林瑛. 利用ELP-Intein系统在大肠杆菌中生产抗菌肽DLP4[J]. Chinese Journal of Biotechnology, 2022, 38(6): 2365-2376

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History
  • Received:November 11,2021
  • Revised:
  • Adopted:
  • Online: June 28,2022
  • Published: June 25,2022
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