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Serial Expression and Activity Analysis of LNK-16: A Bovine Antimicrobial Peptide Analogue

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Abstract

Indolicidin is a broad-spectrum antimicrobial peptide (AMP) with great therapeutic potential; however, high manufacturing costs associated with industrial-scale chemical synthesis have limited its delivery. Therefore, the use of recombinant DNA technology to produce this peptide is urgently needed. In this study, a new methodology for the large-scale production of a novel bovine AMP was developed. LNK-16 is an analogue of indolicidin that contains a kallikrein protease site at its C-terminus. The amino acid sequence of LNK-16 was synthesized using Escherichia coli-preferred codons. Three copies of the target gene were assembled in series by overlapping PCR and cloned into pET-30a(+) for the expression of His-(LNK-16)3 in E. coli BL21 (DE3) cells. The expressed fusion protein His-(LNK-16)3 was purified by Ni2+-chelating chromatography and then cleaved by kallikrein to release LNK-16. The recombinant LNK-16 peptide showed antimicrobial activity similar to that of chemically synthesized LNK-16 and indolicidin. Together, these data indicate that the use of serial expression can improve the large-scale production of AMPs for clinical and research applications.

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Abbreviations

AMPs:

Antimicrobial peptides

G−:

Gram-negative bacteria

G+:

Gram-positive bacteria

MIC:

Minimal inhibitory concentration

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Acknowledgments

This study was supported by National Natural Science Foundation (Grant No. 31372469) and the Science and Technology Innovation Foundation for Distinguished Young Scientists of Henan Province (Grant No. 104100510028).

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Authors and Affiliations

  1. Department of Animal Science and Veterinary Medicine, Henan Institute of Science and Technology, Xinxiang, 453003, Henan, China

    Yanzhao Xu, Qing Wang, Bolin Hang, Dengfeng Fu, Tiantian Shang, Zhiyu Zhao, Qinghua Zhang & Jian-He Hu

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  1. Yanzhao Xu
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  2. Qing Wang
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  3. Bolin Hang
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  4. Dengfeng Fu
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  7. Qinghua Zhang
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  8. Jian-He Hu
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Correspondence to Jian-He Hu.

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Xu, Y., Wang, Q., Hang, B. et al. Serial Expression and Activity Analysis of LNK-16: A Bovine Antimicrobial Peptide Analogue. Protein J 33, 309–312 (2014). https://doi.org/10.1007/s10930-014-9563-0

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