Cloning and Expression of Antibacterial Goat Lactoferricin from Escherichia coli AD494(DE3)pLysS Expression System

Goat lactoferricin (GLfcin), an antibacterial peptide, is released from the N terminus of goat lactoferrin by pepsin digestion. Two GLfcin-related cDNAs, GLfcin L and GLfcin S, encoding Ala20-Ser60 and Ser36-Ser60 of goat lactoferrin, respectively, were cloned into the pET-23a(+) expression vector u...

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Bibliographic Details
Published in:Journal of food protection 2008-12, Vol.71 (12), p.2523-2525
Main Authors: Chen, Gen-Hung, Yin, Li-Jung, Chiang, I-Hua, Jiang, Shann-Tzong
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animal productions
Animals
Anti-Bacterial Agents - pharmacology
Biological and medical sciences
Cloning, Molecular
Consumer Product Safety
DNA, Complementary - analysis
Escherichia coli - genetics
Escherichia coli - metabolism
Food industries
Food microbiology
Fundamental and applied biological sciences. Psychology
Gene Expression
Genetic Vectors - genetics
Goats
Histidine
Humans
Lactoferrin - pharmacology
Recombinant Fusion Proteins - pharmacology
Sequence Analysis, DNA
Temperature
Terrestrial animal productions
Time Factors
Vertebrates
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Summary:Goat lactoferricin (GLfcin), an antibacterial peptide, is released from the N terminus of goat lactoferrin by pepsin digestion. Two GLfcin-related cDNAs, GLfcin L and GLfcin S, encoding Ala20-Ser60 and Ser36-Ser60 of goat lactoferrin, respectively, were cloned into the pET-23a(+) expression vector upstream from (His)6-Tag gene and transformed into Escherichia coli AD494(DE3)pLysS expression host. After being induced by isopropyl-β-D-thiogalactopyranoside (IPTG), two (His)6-Tag fused recombinant lactoferricins, GLfcin L-His·Tag and GLfcin S-His·Tag, were expressed in soluble form within the E. coli cytoplasm. The GLfcin L-His·Tag and GLfcin S-His·Tag were purified using HisTrap affinity chromatography. According to an antibacterial activity assay using the agar diffusion method, GLfcin L-His·Tag had antibacterial activity against E. coli BCRC 11549, Staphylococcus aureus BCRC 25923, and Propionibacterium acnes BCRC 10723, while GLfcin S-His·Tag was able to inhibit the growth of E. coli BCRC 11549 and P. acnes BCRC 10723. These two recombinant lactoferricins behaved as thermostable peptides, which could retain their activity for up to 30 min of exposure at 100°C.
ISSN:0362-028X
1944-9097
DOI:10.4315/0362-028X-71.12.2523