Abstract
Porcine β defensin 2(pBD2)is a cationic 37-amino acid antimicrobial peptide with disulfide bonds. Synthetic pBD2 had broad antimicrobial activity against pathogenic bacteria, and thus pBD2 could be a good candidate as a bactericidal agent for pigs. This study reported the successful recombinant expression of pBD2 in Escherichia coli and analysis of its antimicrobial activity, its hemolytic activity, salt-tolerance and thermal stability as well. The pBD2 gene, obtained by RT-PCR using the tongue total RNA as a template and cloned into pET30a expression vector, was transformed into E. coli BL21 (DE3) plysS. The recombinant pBD2 was expressed after induction by IPTG and purified by His tag affinity column with 95% purity. The recombinant pBD2 exhibited antimicrobial activity against both Gram-positive S. aureus and Gram-negative E. coli including the multi-resistant E. coli. The minimum inhibitory concentration (MIC) of recombinant pBD2 against tested bacteria was 10 μg/mL, and the recombinant pBD2 could kill 50% E. coli at 14.39 μg/mL and S. aureus at 21.1 μg/mL. In addition, pBD2 showed low hemolytic activity, salt-tolerance and thermal stability, the properties would be important for its application in practice.
Keywords: Antimicrobial activity analysis, Porcine β defensin 2, Recombinant expression.
Protein & Peptide Letters
Title:Molecular Cloning, Expression and Characterization of the Porcine β Defensin 2 in E. coli
Volume: 20 Issue: 6
Author(s): Chun-li Li, Yan-cong Zhao, Xiao-yan Song, Xian-xian Huang and Wei-dong Zhao
Affiliation:
Keywords: Antimicrobial activity analysis, Porcine β defensin 2, Recombinant expression.
Abstract: Porcine β defensin 2(pBD2)is a cationic 37-amino acid antimicrobial peptide with disulfide bonds. Synthetic pBD2 had broad antimicrobial activity against pathogenic bacteria, and thus pBD2 could be a good candidate as a bactericidal agent for pigs. This study reported the successful recombinant expression of pBD2 in Escherichia coli and analysis of its antimicrobial activity, its hemolytic activity, salt-tolerance and thermal stability as well. The pBD2 gene, obtained by RT-PCR using the tongue total RNA as a template and cloned into pET30a expression vector, was transformed into E. coli BL21 (DE3) plysS. The recombinant pBD2 was expressed after induction by IPTG and purified by His tag affinity column with 95% purity. The recombinant pBD2 exhibited antimicrobial activity against both Gram-positive S. aureus and Gram-negative E. coli including the multi-resistant E. coli. The minimum inhibitory concentration (MIC) of recombinant pBD2 against tested bacteria was 10 μg/mL, and the recombinant pBD2 could kill 50% E. coli at 14.39 μg/mL and S. aureus at 21.1 μg/mL. In addition, pBD2 showed low hemolytic activity, salt-tolerance and thermal stability, the properties would be important for its application in practice.
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Cite this article as:
Li Chun-li, Zhao Yan-cong, Song Xiao-yan, Huang Xian-xian and Zhao Wei-dong, Molecular Cloning, Expression and Characterization of the Porcine β Defensin 2 in E. coli, Protein & Peptide Letters 2013; 20 (6) . https://dx.doi.org/10.2174/0929866511320060010
DOI https://dx.doi.org/10.2174/0929866511320060010 |
Print ISSN 0929-8665 |
Publisher Name Bentham Science Publisher |
Online ISSN 1875-5305 |
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