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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
 
Creator Lu, Xuan
Hu, Shimeng
Li, Qiaohong
Song, Xia
Zhou, Li
Wang, Yefu
 
Subject Antimicrobial peptide
fusion expression
small ubiquitin-related modifier (SUMO)
subtilosin A
 
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.
 
Date 2022-06-07T11:01:12Z
2022-06-07T11:01:12Z
2021-04
 
Type Article
 
Identifier 0975-0967 (Online); 0972-5849 (Print)
http://nopr.niscpr.res.in/handle/123456789/59869
 
Language en
 
Publisher NIScPR-CSIR, India
 
Source IJBT Vol.20(2) [April 2021]