Antimicrobial Peptide of Korean Native Goat Lactoferrin and Identification of the Part Essential for This Activity

Kimura, Michico; Nam, Myoung Soo; Ohkouchi, Yoshifumi; Kumura, Haruto; Shimazaki, K.; Yu, Dae Yeul

doi:10.1006/bbrc.2000.2141

Cited by 18 publications

(14 citation statements)

References 15 publications

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“…Growth assays were conducted at 37°C (for Staphylococci , Streptococci , Enterococci , K. pneumoniae and yeast‐like fungi), or 27°C (for P. zopfii ) for 24 h. As the growth rate of each microorganism was different, viable cell counts of the stock culture of each microorganism were determined in preliminary studies using standard plate count agar. Based on the results of these preliminary studies, the stock culture of each microorganism was adjusted to 1 × 10 5 cfu/2 μ l. Antimicrobial activity was estimated turbidimetrically by microplate assay (Kimura et al., 2000) using 96‐well microplates and a microplate reader. Approximately 50 μ l of each sterilized culture solution was added to wells of the microplate followed by addition of filter‐sterilized LFH at a concentration of 1 μ g/ml to 70 mg/ml.…”

Section: Methodsmentioning

confidence: 99%

Antibacterial Activity of Bovine Lactoferrin Hydrolysate against Mastitis Pathogens and Its Effect on Superoxide Production of Bovine Neutrophils

Kawai

Shimazaki

Higuchi

et al. 2007

Zoonoses and Public Health

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Antibacterial activity of bovine lactoferrin hydrolysates (LFH) on microorganisms isolated from bovine mastitis, and superoxide (O(2)(-)) production of bovine neutrophils were evaluated. Antibacterial effects of LFH were measured in vitro against Staphylococcus aureus, coagulase-negative staphylococci, Streptococci, Enterococci, Escherichia coli, Klebsiella pneumoniae, yeast-like fungi and Prototheca zopfii isolated from clinical cases of bovine mastitis. To compare susceptibilities against LFH, minimal inhibitory concentration (MIC) values were determined by a micro-plate assay method. Most organisms were sensitive to LFH. Prototheca zopfii was highly sensitive to LFH; the growth of the microorganism was inhibited completely even at 1 mug/ml. Staphylococcus aureus and Escherichia coli were resistant to LFH. The production of O(2)(-) by bovine neutrophils was used to evaluate the effect of LFH administration on functional activity. Increase in O(2)(-) production by bovine neutrophils occurred upon addition of LFH to neutrophils. These results demonstrate that LFH possesses antibacterial activity against pathogens that cause mastitis and activates neutrophil superoxide production.

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Section: Methodsmentioning

confidence: 99%

Antibacterial Activity of Bovine Lactoferrin Hydrolysate against Mastitis Pathogens and Its Effect on Superoxide Production of Bovine Neutrophils

Kawai

Shimazaki

Higuchi

et al. 2007

Zoonoses and Public Health

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“…It was demonstrated that the N-terminal region of lactoferrin is important to the antibacterial activity of lactoferrin (Kang et al, 1996;Kimura et al, 2000). Due to the susceptibility of the lactoferrin N-lobe to proteolytic enzymes (Shimazaki et al, 1993), it is difficult to separate the Nlobe from native lactoferrin.…”

Section: Discussionmentioning

confidence: 99%

Expression of recombinant porcine lactoferrin N-lobe in <i>Pichia</i> <i>methanolica</i> and its antibacterial activity

Shan¹,

Wang²,

Liu³

et al. 2007

J. Anim. Feed Sci.

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Lactoferrin is a multifunctional, iron-binding glycoprotein found in physiological fluids of mammals. Many functions of lactoferrin are thought to be involved in the N-lobe. In the present study, a gene encoding the N-lobe of porcine lactoferrin was cloned and expressed in Pichia methanolica. The gene contains 1038 bp, an "AGG" codon between 1039-1041bp in the PLF gene was replaced with a "TAA" stop codon through primer design and PCR. The sequencing results were aligned with the sequence (Genbank, L77887), indicating that the homology between these two sequences was 99%. The porcine lactoferrin N-lobe (PLfN) was expressed successfully in Pichia methanolica, and was secreted into the culture medium. The protein translated from the new ORF contained 346 amino acid residues, including a 17-amino acid signal peptide, its MW was approximately 42 kD, and it possessed antibacterial activity toward Escherichia coli ATCC25922. The method established in our study will pave the way for efficient industrial production of recombinant PLfN on a large scale for future utilization of this protein as a feed additive.

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“…Antimicrobial activity was measured by 96-well microplate assay (Kimura et al 2000) using P. zopfii (ATCC15627) as the test strain. The test strain was preincubated in heart infusion broth at 27°C.…”

Section: Measurement Of Antimicrobial Activitymentioning

confidence: 99%

Expression of bovine lactoferrin and lactoferrin N-lobe by recombinant baculovirus and its antimicrobial activity against Prototheca zopfii

Tanaka¹,

Nakamura²,

Lee³

et al. 2003

Biochem. Cell Biol.

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Lactoferrin (LF) is a multifunctional, iron-binding glycoprotein found in secretory fluids of mammals. In this study, DNA encoding bovine lactoferrin (bLF) or the N-terminal half of bLF (bLF N-lobe) was inserted into a baculovirus transfer vector, and a recombinant virus expressing bLF or bLF N-lobe was isolated. An 80-kDa bLF-related protein expressed by the recombinant baculovirus was detected by monoclonal antibodies against bLF N-lobe and the C-terminal half of bLF (bLF C-lobe). A 43-kDa bLF N-lobe-related protein expressed by the recombinant baculovirus was detected by anti-bLF N-lobe monoclonal antibody, but not by anti-bLF C-lobe monoclonal antibody. These proteins were also secreted into the supernatant of insect cell cultures. Recombinant bLF (rbLF) and bLF N-lobe (rbLF N-lobe) were affected by tunicamycin treatment, indicating that rbLF and rbLF N-lobe contain an N-linked glycosylation site. Antimicrobial activity of these recombinant proteins against Prototheca zopfii (a yeast-like fungus that causes bovine mastitis) was evaluated by measuring the optical density of the culture microplate. Prototheca zopfii was sensitive to rbLF and rbLF N-lobe, as well as native bLF. There was no difference in antimicrobial activity between rbLF N-lobe and bLF C-lobe.

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Antimicrobial Peptide of Korean Native Goat Lactoferrin and Identification of the Part Essential for This Activity

Cited by 18 publications

References 15 publications

Antibacterial Activity of Bovine Lactoferrin Hydrolysate against Mastitis Pathogens and Its Effect on Superoxide Production of Bovine Neutrophils

Antibacterial Activity of Bovine Lactoferrin Hydrolysate against Mastitis Pathogens and Its Effect on Superoxide Production of Bovine Neutrophils

Expression of recombinant porcine lactoferrin N-lobe in <i>Pichia</i> <i>methanolica</i> and its antibacterial activity

Expression of bovine lactoferrin and lactoferrin N-lobe by recombinant baculovirus and its antimicrobial activity against Prototheca zopfii

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