High Level Expression and Purification of the Clinically Active Antimicrobial Peptide P-113 in Escherichia coli

被引:17
作者
Cheng, Kuang-Ting [1 ,2 ]
Wu, Chih-Lung [1 ,2 ]
Yip, Bak-Sau [1 ,2 ,3 ]
Yu, Hui-Yuan [1 ,2 ]
Cheng, Hsi-Tsung [1 ,2 ]
Chih, Ya-Han [1 ,2 ]
Cheng, Jya-Wei [1 ,2 ]
机构
[1] Natl Tsing Hua Univ, Inst Biotechnol, Hsinchu 300, Taiwan
[2] Natl Tsing Hua Univ, Dept Med Sci, Hsinchu 300, Taiwan
[3] Natl Taiwan Univ Hosp, Dept Neurol, Hsinchu 300, Taiwan
来源
MOLECULES | 2018年 / 23卷 / 04期
关键词
P-113; hG31P; expression; purification; NMR; HISTATIN-CONTAINING MOUTHRINSE; EXPERIMENTAL GINGIVITIS; RECOMBINANT PRODUCTION; ANTIBACTERIAL PEPTIDE; MICROBIAL EVALUATION; FUSION PARTNER; HUMANS;
D O I
10.3390/molecules23040800
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
P-113, which was originally derived from the human saliva protein histatin 5, is a histidine-rich antimicrobial peptide with the sequence AKRHHGYKRKFH. P-113 is currently undergoing phase II clinical trial as a pharmaceutical agent to fight against fungal infections in HIV patients with oral candidiasis. Previously, we developed a new procedure for the high-yield expression and purification of hG31P, an analogue and antagonist of human CXCL8. Moreover, we have successfully removed lipopolysaccharide (LPS, endotoxin) associated with hG31P in the expression with Escherichia coli. In this paper, we have used hG31P as a novel fusion protein for the expression and purification of P-113. The purity of the expressed P-113 is more than 95% and the yield is 4 mg P-113 per liter of E. coli cell culture in Luria-Bertani (LB) medium. The antimicrobial activity of the purified P-113 was tested. Furthermore, we used circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy to study the structural properties of P-113. Our results indicate that using hG31P as a fusion protein to obtain large quantities of P-113 is feasible and is easy to scale up for commercial production. An effective way of producing enough P-113 for future clinical studies is evident in this study.
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页数:11
相关论文
共 33 条
  • [1] Molecular cloning and expression of ranalexin, a bioactive antimicrobial peptide from Rana catesbeiana in Escherichia coli and assessments of its biological activities
    Abou Aleinein, Rasha
    Hamoud, Razan
    Schaefer, Holger
    Wink, Michael
    [J]. APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 2013, 97 (08) : 3535 - 3543
  • [2] Andersson L, 2000, BIOPOLYMERS, V55, P227, DOI 10.1002/1097-0282(2000)55:3<227::AID-BIP50>3.0.CO
  • [3] 2-7
  • [4] Andreu D, 1998, BIOPOLYMERS, V47, P415, DOI 10.1002/(SICI)1097-0282(1998)47:6<415::AID-BIP2>3.0.CO
  • [5] 2-D
  • [6] A new protocol for high-yield purification of recombinant human CXCL8(3-72)K11R/G31P expressed in Escherichia coli
    Cheng, Hsi-Tsung
    Huang, Kuo-Chun
    Yu, Hui-Yuan
    Gao, Kun-Jhih
    Zhao, Xixing
    Li, Fang
    Town, Jennifer
    Gordon, John R.
    Cheng, Jya-Wei
    [J]. PROTEIN EXPRESSION AND PURIFICATION, 2008, 61 (01) : 65 - 72
  • [7] Recombinant expression, purification, and antimicrobial activity of a novel hybrid antimicrobial peptide LFT33
    Feng, Xingjun
    Liu, Chunlong
    Guo, Jiayin
    Song, Xueying
    Li, Jing
    Xu, Wenshan
    Li, Zhongqiu
    [J]. APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 2012, 95 (05) : 1191 - 1198
  • [8] Antimicrobial peptides stage a comeback
    Fox, Jeffrey L.
    [J]. NATURE BIOTECHNOLOGY, 2013, 31 (05) : 379 - 382
  • [9] Antimicrobial and host-defense peptides as new anti-infective therapeutic strategies
    Hancock, Robert E. W.
    Sahl, Hans-Georg
    [J]. NATURE BIOTECHNOLOGY, 2006, 24 (12) : 1551 - 1557
  • [10] Production in Escherichia coli of moricin, a novel type antibacterial peptide from the silkworm, Bombyx mori
    Hara, S
    Yamakawa, M
    [J]. BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 1996, 220 (03) : 664 - 669