Expression of Enterocin-P in HEK Platform: Evaluation of Its Cytotoxic Effects on Cancer Cell Lines and Its Potency to Interact with Cell-Surface Glycosaminoglycan by Molecular Modeling

Pirkhezranian, Zana; Tanhaeian, Abbas; Mirzaii, Mehdi; Sekhavati, Mohammad Hadi

doi:10.1007/s10989-019-09956-7

Cited by 13 publications

(8 citation statements)

References 34 publications

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“…The chimeric lactoferricin and lactoferrampin consists of 36 amino acids and was generated through the fusion of two short regions of camel lactoferrin 284 DLIWKLLVKAQEKFGRGKPS 303 (ID: AHJ37525) and 49 RVKKMRRQWQACKSS 35 (ID:NP_001290496.1) linked by Lysine (GenBank accession number: MH327768). The chimeric peptide encoding sequence was codon optimized for the appropriate expression in L. lactis by Genscript® (USA).…”

Section: Gene Synthesis and Vector Constructionmentioning

confidence: 99%

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Generation of an engineered food-grade Lactococcus lactis strain for production of an antimicrobial peptide: in vitro and in silico evaluation

et al. 2020

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Background: Foodborne pathogens and their biofilms are considered as one of the most serious problems in human health and food industry. Moreover, safety of foods is a main global concern because of the increasing use of chemical food additives. Ensuring food safety enhances interest in discovery of new alternative compounds such as antimicrobial peptides (AMPs), which can be used as bio-preservatives in the food industry. In this study, the most important antimicrobial peptides of camel milk lactoferrin (lactoferrampin and lactoferricin) were recombinantly expressed in the form of chimeric peptide (cLFchimera) in a food-grade L. lactis strain. P170 expression system was used to express secreted cLFchimera using pAMJ1653 expression vector which harbors a safe (non-antibiotic) selectable marker. Results: Peptide purification was carried out using Ni-NTA agarose column from culture medium with concentration of 0.13 mg/mL. The results of disk diffusion test revealed that cLFchimera had considerable antimicrobial activity against a number of major foodborne bacteria. Furthermore, this chimeric peptide showed strong and weak inhibitory effect on biofilm formation against P. aeruginosa, S. aureus E. faecalis, and E. coli, respectively. Antioxidant activity and thermal stability of the chimeric peptide was determined. The results showed that cLFchimera had antioxidant activity (IC 50 : 310 μ/mL) and its activity was not affected after 40 min of boiling. Finally, we evaluated the interaction of the peptide with LPS and DNA in bacteria using molecular dynamic simulation as two main intra and extra cellular targets for AMPs, respectively. Our in silico analysis showed that cLFchimera had strong affinity to both of these targets by positive charged residues after 50 ns molecular dynamic simulation. Conclusions: Overall, the engineered food-grade L. lactis generated in the present study successfully expressed a secreted chimeric peptide with antimicrobial properties and could be considered as a promising bio-preservative in the food industry.

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Section: Gene Synthesis and Vector Constructionmentioning

confidence: 99%

“…AMP has been known as a natural molecule with activity toward a broad spectrum of microorganisms including bacteria, fungi, and viruses [11]. Moreover, AMPs have been considered as a new generation of biologically active regulators that can prevent oxidation and microbial degradation in foods [35,49].…”

Section: Introductionmentioning

confidence: 99%

Generation of an engineered food-grade Lactococcus lactis strain for production of an antimicrobial peptide: in vitro and in silico evaluation

et al. 2020

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“…AMPs play an important role in natural defense mechanism for destroying microbial infections [2]. AMPs have a net positive charge and an amphipathic structure and usually contain 12-50 amino acid residues [3,4]. More recently, a chimeric form of peptide named cLFchimera has been expressed and puri ed in E. coli [5] in our lab.…”

Section: Introductionmentioning

confidence: 99%

Antibacterial Activity of cLFchimera and its Synergistic Potential with Antibiotics against Some Foodborne Pathogens Bacteria

Roshanak

Pirkhezranian

Shahidi

et al. 2020

Preprint

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Background: Frequent and unlimited use of antibiotics caused the development of antibiotic resistance by microorganisms. Therefore, there is an argent need to discover novel antibacterial agents or a combination of agents as a safe treatment strategy for various infections. The aim of the present study was to evaluate the synergistic effects of cLFchimera, an antimicrobial peptides (AMPs), and antibiotics on several foodborne bacterial strains. Methods: A checkerboard method was used to determine the synergistic effects of cLFchimera and several antibiotics (Gentamicin, Cefazolin and Ceftazidime) on bacterial strains (Escherichia coli, Pseudomonas aeruginosa and Salmonella typhi). Results: The combination of cLFchimera and antibiotics generated a total and partial synergistic interaction for all foodborne bacterial strain used in the present study (FIC= 0.25 to 0.77). In most cases, the effect of peptide and antibiotic synergist on release of cellular content was not different compared to antibiotics when they used alone, but the count of viable cells significantly decreased in combination peptide and antibiotics treatments. Generally, antibacterial dynamics of the combination of peptide and antibiotics showed an increase and stable trend after reaching the peak point for E. coli, P. aeruginosa and S. typhi, respectively. Scanning electron microscopy analysis showed that bacterial cells treated with the combination Gentamycin and cLFchimera were markedly damaged, and most of the outermost layer of the bacterial cells disappeared. Conclusion: Overall, our results may suggest that cLFchimera mediated its synergistic activity independent to antibiotics mode of action by disrupting the cell membrane and intramolecular mechanisms.

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“…Antimicrobial peptides (AMPs) are endo-exogenous polypeptides comprised of less than 50 amino acids, characterized by cationic amphipathic properties, and produced by host defense systems or synthetically supplied to the diet in order to protect a host from pathogenic microbes [5, 6]. AMPs show broad-spectrum antimicrobial activities against various microorganisms, including Gram-positive and Gram-negative bacteria, fungi, and viruses [5].…”

Section: Introductionmentioning

confidence: 99%

Effects of cLFchimera, a recombinant antimicrobial peptide, on intestinal morphology, microbiota, and gene expression of immune cells and tight junctions in broiler chickens challenged withC. perfringens

Daneshmand

Kermanshahi

et al. 2019

Preprint

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24The current study was conducted to investigate the effects of cLFchimera, a recombinant 25 antimicrobial peptide (AMP), on various productive performance and gut health attributes of 26 broilers experimentally challenged with Clostridium perfringens (Cp). Three hundred and sixty 1-27 day-old chickens were randomly allocated to 4 treatments of 6 replicates as follows: T1) 28 unchallenged group fed with corn-soybean meal (CSM) without Cp challenge and additives; T2) 29 challenge group fed with CSM and challenged with Cp without any additives; T3) peptide group 30 challenged with NE supplemented with 20 mg cLF36/kg diet (AMP); T4) antibiotic group 31 challenged with NE and supplemented with 45 mg antibiotic (bacitracin methylene disalicylate)/kg 32 diet (antibiotic). Birds had free access to feed and water, sampling for villi morphology and ileal 33 microbiota were performed on days 10 and 22, while jejunal section was sampled for gene 34 expression of cytokines, tight junctions proteins, and mucin only on day 22. Results showed that 35 AMP ameliorated NE-related lesion in the jejunum and ileum and reduced mortality in challenged 36 birds compared to challenge group with Cp without any additives. Also, supplementing challenged 37 birds with AMP improved growth performance and reconstructed villi morphology. While 38 antibiotic non-selectively reduced the count of bacteria, AMP positively restored ileal microflora 39 in favor of good bacteria (i.e. Bifidobacteria spp. and Lactobacillus spp.). AMP beneficially 40 regulated the expression of cytokines, junctional proteins, and mucin in the jejunum of challenged 41 birds with Cp. Since cLFchimera ameliorated NE lesion score, reduced mortality, improved 42 productive performance and gut health attributes in chickens compared to challenged group and 43 also were mostly similar with those of antibiotics and therefore, it could be concluded that this 44 chimeric peptide can be a worthy candidate to substitute growth promoter antibiotics, while more 45 research is required to unveil the exact mode of action of this synthetic peptide. 46 47 Author summary 48 Necrotic enteritis (NE) is a detrimental enteric disease in the poultry industry worldwide. The 49 etiological factor of this disease is Clostridium perfringens, which is gram-positive anaerobic 50 bacterium. This bacterium is common inhabitant of the intestine in lower counts (105), but it 51 becomes pathogenic in higher counts and can secrete NetB toxin, which is the main cause of 52 inducing NE in broilers. Due to the emergence of antibiotic-resistant bacteria, new generation of 53 antimicrobial additives such as antimicrobial peptides (AMPs) has been introduced to the poultry 54 industry. AMPs are small molecules with 12-50 amino acids having antibacterial activity. 55 Recently, we extracted new AMP from camel milk, expressed in E. coli, refined and lyophilized 56 to produce purified peptides. The current study investigated the effects of this peptide on 57 prevention of NE in broilers. Results showed th...

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Expression of Enterocin-P in HEK Platform: Evaluation of Its Cytotoxic Effects on Cancer Cell Lines and Its Potency to Interact with Cell-Surface Glycosaminoglycan by Molecular Modeling

Cited by 13 publications

References 34 publications

Generation of an engineered food-grade Lactococcus lactis strain for production of an antimicrobial peptide: in vitro and in silico evaluation

Generation of an engineered food-grade Lactococcus lactis strain for production of an antimicrobial peptide: in vitro and in silico evaluation

Antibacterial Activity of cLFchimera and its Synergistic Potential with Antibiotics against Some Foodborne Pathogens Bacteria

Effects of cLFchimera, a recombinant antimicrobial peptide, on intestinal morphology, microbiota, and gene expression of immune cells and tight junctions in broiler chickens challenged withC. perfringens

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