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In vitro reconstitution of antimicrobial pathogen activity by expressed recombinant bovine lactoferrin N-terminal peptide in Escherichia coli

Published online by Cambridge University Press:  30 April 2007

Hongxia Luo ,
Shangwu Chen ,
Fazheng Ren ,
Huiyuan Guo ,
Shaohua Lin  and
Wentao Xu
Hongxia Luo
Affiliation:
Key Lab of Functional Dairy Science of Beijing and Ministry of Education of China, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, China 100083 Beijing Vocational College of Agriculture, Beijing, China 102442
Shangwu Chen
Affiliation:
Key Lab of Functional Dairy Science of Beijing and Ministry of Education of China, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, China 100083
Fazheng Ren
Affiliation:
Key Lab of Functional Dairy Science of Beijing and Ministry of Education of China, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, China 100083
Huiyuan Guo
Affiliation:
Key Lab of Functional Dairy Science of Beijing and Ministry of Education of China, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, China 100083
Shaohua Lin
Affiliation:
Key Lab of Functional Dairy Science of Beijing and Ministry of Education of China, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, China 100083
Wentao Xu
Affiliation:
Key Lab of Functional Dairy Science of Beijing and Ministry of Education of China, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, China 100083
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Abstract

Recombinant bovine lactoferrin N-terminal polypeptide (rbLF-N) Escherichia coli expression system was constructed and the rbLF-N antimicrobial activity was displayed by enzymatic proteolysis in this study. A 162 bp 5′-terminal fragment of bovine lactoferrin (bLF) gene from bovine liver gDNA was amplified by PCR. The DNA fragment containing exon-2 of the bLF gene was cloned into the expression vector pGEX-4T1 and the glutathione-S-transferase–rbLF-N (GST-rbLF-N) fusion protein was obtained by over-expression in Esch. coli BL21(DE3). After thrombin/pepsin digestion, the rbLF-N was released from the fusion protein. The recombinant peptide was separated and identified by SDS-PAGE, HPLC and LC-MS/MS analysis. A very strong anti-food-born microbial pathogen activity of the rbLF-N peptides was displayed through bio- and kinetic-assays in vitro. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the rbLF-N peptide for bacterial pathogens Staphylococcus aureus, Streptococcus mutans, Esch. coli and Klebsiella pneumoniae were 11·7, 11·7, 11·7, 23·4 μg and 23·4, 11·7, 11·7, 46·4 μg, respectively. This study created a new route for exploring lactoferrin peptide application in food science.

Keywords

Lactoferrinanti-bacterial pathogen peptiderbLF-NEscherichia coli
Type
Research Article
Information
Journal of Dairy Research , Volume 74 , Issue 2 , May 2007 , pp. 233 - 238
Copyright
Proprietors of Journal of Dairy Research 2007

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