High-yield soluble expression and simple purification of the antimicrobial peptide OG2 using the intein system in Escherichia coli

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DOIResolve DOI: http://doi.org/10.1155/2013/754319
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TypeArticle
Journal titleBioMed Research International
ISSN2314-6133
Volume2013
Article number754319
Subjectbacterial protein; intein; og 2 protein; polypeptide antibiotic agent; unclassified drug; antimicrobial activity; article; Escherichia coli; molecular cloning; nonhuman; protein expression; protein purification
AbstractOG2 is a modified antimicrobial peptide, that is, derived from the frog peptide Palustrin-OG1. It has high antimicrobial activity and low cytotoxicity, and it is therefore promising as a therapeutic agent. Both prokaryotic (Escherichia coli) and eukaryotic (Pichia pastoris) production host systems were used to produce OG2 in our previous study; however, it was difficult to achieve high expression yields and efficient purification. In this study, we achieved high-yield OG2 expression using the intein fusion system. The optimized OG2 gene was cloned into the pTWIN1 vector to generate pTWIN-OG2-intein2 (C-terminal fusion vector) and pTWIN-intein1-OG2 (N-terminal fusion vector). Nearly 70% of the expressed OG2-intein2 was soluble after the IPTG concentration and induction temperature were decreased, whereas only 42% of the expressed of intein1-OG2 was soluble. Up to 75 mg of OG2-intein2 was obtained from a 1 l culture, and 85% of the protein was cleaved by 100 mM DTT. Intein1-OG2 was less amenable to cleavage due to the inhibition of cleavage by the N-terminal amino acid of OG2. The purified OG2 exhibited strong antimicrobial activity against E. coli K88. The intein system is the best currently available system for the cost-effective production of OG2. © 2013 Yong-Gang Xie et al.
Publication date
LanguageEnglish
AffiliationNational Research Council Canada (NRC-CNRC); NRC Biotechnology Research Institute (BRI-IRB)
Peer reviewedYes
NPARC number21269652
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Record identifier7cd1df62-bf19-4b8f-b8d7-26e00c8d6876
Record created2013-12-13
Record modified2016-05-09
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