Effects of Thioredoxin: SUMO and Intein on Soluble Fusion Expression of an Antimicrobial Peptide OG2 in Escherichia coli

Author(s): Yong-Gang Xie, Chao Luan, Hai-Wen Zhang, Fei-Fei Han, Jie Feng, Young-Jun Choi, Denis Groleau, Yi-Zhen Wang.

Journal Name: Protein & Peptide Letters

Volume 20 , Issue 1 , 2013

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OG2 is a modified antimicrobial peptide of Palustrin-OG1 (OG1), which is derived from Odorrana grahami frog. OG2 has shown much higher selective antimicrobial activity and lower hemolytic activity than OG1, indicating OG2 may be a promising antimicrobial agent. In this study, we investigated three fusion partners, including thioredoxin, Mxe GyrA intein, and small ubiquitin-like modifier (SUMO), each fused with OG2, and examined their effects on the expression level and solubility of OG2 in Escherichia coli. The codon-optimized OG2 gene was cloned into pET32a (+) and pTWIN1 for fusion with thioredoxin and Mxe GyrA intein, respectively. In addition, the SUMO-OG2 gene was amplified by splice overlap extension PCR method and was cloned into pET30a (+). All recombinant plasmids were then transformed into E. coli BL21(DE3)pLysS, and the expressed fusion proteins were verified. Upon isopropyl β-D-1- thiogalactopyranoside (IPTG) induction, OG2 fused with thioredoxin (Trx-OG2) showed the highest yield as a soluble fusion protein (50 mg/L), followed by Mxe GyrA intein (44 mg/L) and SUMO (11 mg/L). The thioredoxin-fused protein (Trx-OG2) was then purified by nickel-nitrilotriacetic acid chromatography and desalted by Sephadex G25. The OG2 released by both tobacco etch virus protease and enterokinase from Trx-OG2 showed strong antimicrobial activity against Staphylococcus aureus ATCC25923.

Keywords: Antimicrobial activity, enterokinase, Escherichia coli, fusion partner, hemolysis, OG2, tobacco etch virus protease, antimicrobial peptide, hemolytic activity, thioredoxin

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Article Details

Year: 2013
Page: [54 - 60]
Pages: 7
DOI: 10.2174/0929866511307010054

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