Production of Recombinant Porcine Lactoferrin Exhibiting Antibacterial Activity in Methylotrophic Yeast, &lt;i&gt;Pichia pastoris&lt;/i&gt;

Chen, Hsiao Ling; Lai, Yi; Yen, Chih-Ching; Lin, Yuh Yih; Lu, Ching-Lan; Yang, Su–Hua; Tsai, Tsung-Neng; Lin, Yi; Lin, Chin Wen; Chen, Chuan-Mu

doi:10.1159/000085787

Cited by 30 publications

(39 citation statements)

References 63 publications

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“…The MIC and minimal bactericidal concentration (MBC) of a pepsin-digested r-pLf hydrolysate against E. coli were 150 and 200 μg/ml, respectively, while intact r-pLf had an MIC of 750 μg/ml. The peptides obtained by pepsin digestion of r-pLf exhibited more antimicrobial activity than native r-pLf, apparently because they disrupt the cell wall and disintegrate the LPS molecules of the outer membrane [77].…”

Section: In Vivo Assaysmentioning

confidence: 98%

Lactoferrin: A Powerful Antimicrobial Protein Present in Milk

Luna-Castro¹,

Samaniego-Barrón²,

Serrano-Rubio³

et al. 2017

J Adv Dairy Res

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Lactoferrin (Lf) is an iron-chelating glycoprotein present in milk and mucosal secretions, a component of the mammalian innate immune system. Lf is microbiostatic and microbicidal. Lf can reduce the bacterial expression of virulence factors, such as those involved in biofilm production and protease secretion. The high identity among mammalian Lf sequences facilitates its use in human and veterinary medicine. Lf of bovine origin is the principal Lf used due to its commercial availability through purification from milk whey; recombinant Lfs (bovine, human, and porcine) have been used as well. Lf is a stable protein that retains its physicochemical characteristics under gastric pH conditions, and in most cases it is bioactive even after digestion; thus, the incorporation of Lf into diets facilitates its administration to animals. The aim of this review is to examine original research in which the effects of bovine and porcine Lf on pathogens of domestic animals have been demonstrated through in vitro and in vivo assays, with the purpose of ascertaining the benefits that Lf provides in the treatment of infectious diseases.

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Section: In Vivo Assaysmentioning

confidence: 98%

Lactoferrin: A Powerful Antimicrobial Protein Present in Milk

Luna-Castro¹,

Samaniego-Barrón²,

Serrano-Rubio³

et al. 2017

J Adv Dairy Res

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“…Therefore, LF is postulated to be involved in the innate host defense against infection and severe inflammation, most notable at mucosal surfaces such as those of the gastrointestinal tract (33). Antimicrobial properties of LF include bacteriostasis by iron deprivation (34), bactericidal activity by destabilization of the cell wall (7,10), and antiviral activity by inhibition of viral infection (35). Besides antimicrobial activity, LF has been shown to promote the growth of Bifidobacterium species, the predominant bacteria of the intestinal flora of healthy breast-fed newborns (36).…”

Section: Discussionmentioning

confidence: 99%

“…The structure and functions of bovine and human LF have been well characterized (13)(14)(15), but little is known about porcine LF. In a previous study, we generated an expression system using a recombinant methylotrophic yeast, Pichia pastoris, which harbored the pLF gene driven by the inducible promoter of the alcohol oxidase 1 gene (AOX1) and the yeast R-mating factor signal peptide (7). Significant levels of secreted pLF were achieved in culture medium of the yeast transformants.…”

Section: Introductionmentioning

confidence: 99%

Recombinant Porcine Lactoferrin Expressed in the Milk of Transgenic Mice Enhances Offspring Growth Performance

Chen

Yen

et al. 2007

J. Agric. Food Chem.

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The European Commission has proposed a permanent ban on the use of antibiotics as an ingredient in animal feed to promote growth. Lactoferrin is a globular multifunctional protein that has been shown to play a role in iron absorption and to have antimicrobial and anti-inflammatory activities. Therefore, lactoferrin may serve as a nontherapeutic alternative to antibiotics in livestock husbandry. As a pilot study toward this goal, transgenic mice have been generated harboring a porcine lactoferrin (pLF) gene driven by the mammary gland-specific promoter of the bovine R-lactalbumin (RLA) gene. The RLA-pLF hybrid gene was confirmed to have been successfully integrated and transmitted stably through the germ-line in 9 (5 females and 4 males) of 14 transgenic founders. In the female progenies of six lines analyzed, the transgene copy numbers ranged from 1 to 20 with 1-4 integration sites. Significant levels of pLF protein in milk ranging from 40 to 106 µg/mL with physical characteristics similar to those of native pLF in sow's milk were achieved in three of the transgenic lines obtained. Tissue-and stage-specific pLF expressions were restricted to the mammary gland of the transgenic female mice during lactation. It was further demonstrated that the growth performance of animal pups is enhanced by directly feeding the genetically engineered milk containing enriched pLF protein in transgenic mice. Furthermore, this enhanced growth performance in suckling mice was proportional to the concentration of pLF present in milk.

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“…Determination of the bacterial inhibition curve of the recombinant pLF protein produced from L. pentosus transformant was performed using a standard protocol with an inoculum of 1 × 10 3 bacteria colony-forming units per ml in MRS medium (for lactobacillus) or Luria-Bertani medium (for other tested bacteria), as described earlier (Chen et al, 2004). The tested bacteria include Staphylococcus aureus strain CVCC26003, Bacillus subtilis strain CVCC63501, Escherichia coli strain CVCC10141, Salmonella enterica ssp.…”

Section: In Vitro Antibacterial Activity Assaysmentioning

confidence: 99%

Lactobacillus pentosus expressing porcine lactoferrin elevates antibacterial activity and improves the efficacy of vaccination against Aujeszky’s disease

Zong

Han

et al. 2016

Acta Veterinaria Hungarica

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In this study, Lactobacillus pentosus expressing porcine lactoferrin (pLF) was tested for in vitro antibacterial activity and for its ability to enhance immunity induced by an orally administered Aujeszky's disease virus (ADV) vaccine. The cDNA encoding N-terminus of pLF was cloned into a Lactobacillus-specific plasmid to produce L. pentosus pLF expressing transformants (pPG612.1-pLF-N/L. pentosus). The antimicrobial activity of the recombinant pLF protein inhibited bacterial growth in vitro. The supernatant of pPG612.1-pLF-N/L. pentosus had an inhibitory effect on Staphylococcus aureus strain CVCC26003, Bacillus subtilis strain CVCC63501, Escherichia coli strain CVCC10141 and Salmonella enterica ssp. enterica Choleraesuis strain CVCC79102, while it did not inhibit the growth of Lactobacillus casei strain ATCC393. A mouse model was established to test the effectiveness of the orally administered probiotic L. pentosus recombinant strain in the gastrointestinal tract. Mice were immunised with an attenuated porcine Aujeszky's disease virus (ADV) vaccine. Serum antibody levels determined using a mouse Aujeszky's disease IgG ELISA showed that IgG levels were significantly higher in the pPG612.1-pLF N /L. pentosus group than in the PBS and Lactobacillus pentosus groups at days 7 and 21 (P < 0.01) and at day 14 (P < 0.05), indicating that this oral recombinant strain can improve the effectiveness of the vaccine and play a role in immune enhancement through humoral immunity. These results suggest that the recombinant Lactobacillus pentosus not only has the beneficial characteristics of lactic acid bacteria but also produces biologically functional lactoferrin.Key words: Porcine lactoferrin, recombinant Lactobacillus pentosus, antibacterial activity, immune enhancement Lactoferrin (LF) is an 80-kDa iron-binding glycoprotein. It is abundantly found in exocrine secretions of mammals and especially in milk and fluids of the digestive tract released by mucosal epithelia and neutrophils during inflamma-

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Production of Recombinant Porcine Lactoferrin Exhibiting Antibacterial Activity in Methylotrophic Yeast, <i>Pichia pastoris</i>

Cited by 30 publications

References 63 publications

Lactoferrin: A Powerful Antimicrobial Protein Present in Milk

Lactoferrin: A Powerful Antimicrobial Protein Present in Milk

Recombinant Porcine Lactoferrin Expressed in the Milk of Transgenic Mice Enhances Offspring Growth Performance

Lactobacillus pentosus expressing porcine lactoferrin elevates antibacterial activity and improves the efficacy of vaccination against Aujeszky’s disease

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