Expression, purification and antimicrobial activity analysis of recombination peptide subtilosin A in Escherichia coli using SUMO fusion technology
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Title |
Expression, purification and antimicrobial activity analysis of recombination peptide subtilosin A in Escherichia coli using SUMO fusion technology
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Creator |
Lu, Xuan
Hu, Shimeng Li, Qiaohong Song, Xia Zhou, Li Wang, Yefu |
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Subject |
Antimicrobial peptide
fusion expression small ubiquitin-related modifier (SUMO) subtilosin A |
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Description |
138-144
Subtilosin A is an antimicrobial peptide isolated from Bacillus subtilis, and it possess broad-spectrum antibacterial activity. As subtilosin A prevents biofilm formation by inhibiting bacterial quorum sensing processes against Gram-positive, Gram-negative, and Gram-variable bacteria, it is urgent to obtain high-quality subtilosin A production in an economical and effective way. Escherichia coli is undoubtedly a most preferred host system for production of heterologous recombinant proteins. However, subtilosin A has the inhibitory effects against E. coli and it is easily to be degraded. To produce subtilosin A in E. coli, the application of small ubiquitin-related modifier (SUMO) fusion system with the purpose of stability and prevention of antimicrobial activity is the best practices in this regard. In this study, subtilosin A gene with codon optimization was cloned into the Stu I/Hind III sites of pSUMO vector after SUMO tag and transformed to E. coli BL21 (DE3). The SUMO-subtilosin A fusion protein was induced at 15ºC for 16 hours with 0.5 mM isopropyl thio-β-D- galactoside (IPTG) induction. The fusion protein with a molecular weight of approximately 20 kDa was confirmed by SDS-PAGE. The subtilosin A was then released by SUMO protease cleavage at the junction site and isolated and purified by affinity and cation exchange chromatography. The recombinant subtilosin A demonstrated a good antibacterial activity against Pseudomonas aeruginosa (Gram-negative bacteria) and Staphylococcus aureus (Gram-positive bacteria), with minimum inhibitory concentrations (MICs) of 100 mg/L and 100 mg/L, respectively. These results showed an efficient method for synthesis functional subtilosin A. |
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Date |
2022-06-07T11:01:12Z
2022-06-07T11:01:12Z 2021-04 |
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Type |
Article
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Identifier |
0975-0967 (Online); 0972-5849 (Print)
http://nopr.niscpr.res.in/handle/123456789/59869 |
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Language |
en
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Publisher |
NIScPR-CSIR, India
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Source |
IJBT Vol.20(2) [April 2021]
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