期刊
PROTEIN EXPRESSION AND PURIFICATION
卷 94, 期 -, 页码 73-78出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.pep.2013.11.001
关键词
Antibacterial activity; Channel catfish; Hepcidin; Recombinant protein expression; Fusion protein; Enterokinase
类别
资金
- Innovation Program of Shanghai Municipal Education Commission [11ZZ148]
- League Program from the Science and Technology Commission of Shanghai Municipality [4319]
Hepcidins are small cysteine-rich cationic antimicrobial peptides. The channel catfish (Ictalurus punctatus) hepcidin cDNA has been characterized, but recombinant protein expression and purification was not reported. I. punctatus hepcidin is comprised of 96 residues, with eight functionally important conserved cysteine residues located in the C-terminal region of the mature peptide, suggesting that this region is responsible for the antibacterial activity. In this study, a cDNA fragment (mCH) encoding the 25 amino acid mature peptide was cloned from channel catfish liver, and inserted into vector pET-32a(+) to produce a construct that expressed a hexahistidine-tagged thioredoxin (trxA) fusion protein that was cleavable using enterokinase. The trxA-mCH fusion protein was expressed in Escherichia coli BL21 (DE3) at 25 degrees C, using 1 mM IPTG for induction. Greater than 80% of the fusion protein was expressed solubly, but was not biologically active. Removal of the trxA fusion partner by enterokinase resulted in mCH that exhibited antibacterial activity against two Gram-positive (Listeria monocytogenes and Staphylococcus aureus), and two Gram-negative (E. coli and Pseudomonas aeruginosa) bacteria. (C) 2013 Elsevier Inc. All rights reserved.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据