Constructing a hybrid molecule with low capacity of IgE binding from Chenopodium album pollen allergens

Nouri, Hamid Reza; Varasteh, Abdolreza; Vahedi, Fatemeh; Chamani, Jamshidkhan; Afsharzadeh, Danial; Sankian, Mojtaba

doi:10.1016/j.imlet.2012.03.008

Cited by 14 publications

(12 citation statements)

References 32 publications

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“…We have already constructed an rHM of three identified C. album pollen allergens, Che a 1, Che a 2 and Che a 3, via overlapping extension PCR. Using immunological experiments, we have confirmed that the constructed rHM has strongly lost its allergenicity and proposed that rHM may act as a suitable option in immunotherapy [16]. Since immunogenic properties of recombinant allergens and hypoallergenic derivatives must be evaluated in vivo before being used in immunotherapy of allergic patients [19], we studied immunotherapeutic potential of constructed rHM on type I allergy.…”

Section: Discussionmentioning

confidence: 99%

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Immunotherapy with a Recombinant Hybrid Molecule Alleviates Allergic Responses More Efficiently than an Allergenic Cocktail or Pollen Extract in a Model of Chenopodium album Allergy

Nouri

Sankian²,

Afsharzadeh³

et al. 2013

Int Arch Allergy Immunol

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Background: The aim of this study is to assess the therapeutic potential of a recombinant hybrid molecule (rHM) alongside an allergenic cocktail from recombinant wild-type allergens as well as pollen extract on Chenopodium album allergy, using a BALB/c mouse model. Methods: The BALB/c mice had already been sensitized to C. album via intraperitoneal injections of alum-adsorbed allergenic cocktail and immunotherapy procedure was followed by subcutaneous injections of the rHM, allergenic cocktail and pollen extract at weekly intervals. Humoral immune responses were determined via measurement of specific antibodies in serum. Splenocytes of immunized mice were stimulated in vitro and then proliferation responses, cytokine secretion and mRNA expression of genes involved in immunotherapy were examined by ELISA and real-time PCR. Results: Sensitized mice were identified with high specific IgE against allergenic cocktail when compared with healthy mice. Immunotherapy with the rHM induced the highest ratio of the IgG2a/IgG1 levels compared to allergenic cocktail or C. album pollen extract. The rHM was able to induce proliferative responses as well as the allergenic cocktail in cultured splenocytes. Immunotherapy with the rHM significantly improved secretion of IFN-γ and IL-10, while secretion of IL-13 rapidly diminished. Interestingly, mRNA expression of GATA3 was strongly decreased in rHM-treated mice whereas mRNA expression of T-bet and Foxp3 was significantly increased. Conclusion: Our results prove that immunotherapy with the rHM effectively controlled allergic responses by shifting from a Th2-like immune response to a Th1-dominated immune response.

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Section: Discussionmentioning

confidence: 99%

“…Immunologic experiments confirmed that the rHM had low capacity of IgE binding while its immunogenicity was preserved [16]. The recombinant Che a 1, Che a 2 and Che a 3 were previously expressed and purified from E. coli [13] and we had built an allergenic cocktail with equimolar concentrations of them.…”

Section: Methodsmentioning

confidence: 99%

Immunotherapy with a Recombinant Hybrid Molecule Alleviates Allergic Responses More Efficiently than an Allergenic Cocktail or Pollen Extract in a Model of Chenopodium album Allergy

Nouri

Sankian²,

Afsharzadeh³

et al. 2013

Int Arch Allergy Immunol

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“…The cDNA coding of the chimeric DNA consists of Che a 1, Che a 2, and Che a 3, three identified allergens of C. album, was constructed using overlapping extension PCR as previously described [16]. The lyophilized E. coli DH5a that containing pET21b?…”

Section: Amplification Of the Chimeric Dna And Preparation Of Expressmentioning

confidence: 99%

“…The resultant supernatant was save as inclusion bodies phase. Purification of the chimeric allergen from soluble and inclusion bodies phases were followed by Ni-NTA chromatography (Qiagen, USA) as previously described [16]. The protein content was determined by Bradford method [4].…”

Section: Transformation Of Recombinant Plasmids Into the E Coli Stramentioning

confidence: 99%

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Expression of a Chimeric Allergen with High Rare Codons Content in Codon Bias-Adjusted Escherichia coli: Escherichia coli BL21 (DE3)-Codon Plus RIL as an Efficient Host

et al. 2016

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The expression of heterologous proteins in Escherichia coli (E. coli) is importantly affected by codon bias. Hence, the aim of the current study was to determine which codon bias-adjusted E. coli strain is sufficient for expression of a chimeric allergen coded by high rare codon content. To investigate the expression level, a chimeric protein of Chenopodium album (C. album) was used as an appropriate model. An expression construct was assembled and was transformed to four strains of codon bias-adjusted E. coli including origami, BL21 (DE3), BL21 (DE3)-codon plus RIL, and Rosetta. The level of expression and solubility of the chimeric allergen was analyzed by SDS-PAGE. In addition, the allergenicity of chimeric allergen was determined using immunoblotting. Our results showed that the chimeric allergen was expressed at high level in E. coli BL21 (DE3)-codon plus RIL and Rosetta. In detail, this recombinant allergen was isolated from soluble fraction in the codon bias-adjusted strains of E. coli BL21 (DE3)-codon plus RIL and Rosetta. Moreover, some lower molecular weight proteins were observed in Rosetta, which could be related to inappropriate expression or broken compartments of the chimeric allergen. The immunoblotting assay confirmed that the IgE-specific immune reactivity of our chimeric allergen expressed in BL21 (DE3)-codon plus RIL was significantly higher than the other strains. Our results showed that the expression of the chimeric allergen with high rare codons content in a codon bias-adjusted strain E. coli BL21 (DE3)-codon plus RIL improves the quality and solubility of the heterologous protein production.

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Genetically engineered fusion of allergen and viral-like particle induces a more effective allergen-specific immune response than a combination of them

Sani

Bargahi

Momenzadeh

et al. 2020

Appl Microbiol Biotechnol

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Chimeric virus-like particles (VLPs) were developed as a candidate for allergen-specific immunotherapy. In this study, hepatitis B core antigen (HBcAg) that genetically fused to Chenopodium album polcalcin (Che a 3)-derived peptide was expressed in E. coli BL21, purified, and VLP formation was evaluated using native agarose gel electrophoresis (NAGE) and transmission electron microscopy (TEM). Chimeric HBc VLPs were characterized in terms of their reactivity to IgE, the induction of blocking IgG and allergen-specific IgE, basophil-activating capacity, and Th1-type immune responses. Results from IgE reactivity and basophil activation test showed that chimeric HBc VLPs lack IgE-binding capacity and basophil degranulation activity. Although chimeric HBc VLPs induced the highest level of efficient polcalcin-specific IgG antibody in comparison to those induced by recombinant Che a 3 (rChe a 3) mixed either with HBc VLPs or alum, they triggered the lowest level of polcalcin-specific IgE in mice following immunization. Furthermore, in comparison to the other antigens, chimeric HBc VLPs produced a polcalcinspecific Th1 cell response. Taken together, genetically fusion of allergen derivatives to HBc VLPs, in comparison to a mix of them, may be a more effective way to induce appropriate immune responses in allergen-specific immunotherapy. Key points• The insertion of allergen-derived peptide into major insertion region (MIR) of hepatitis B virus core (HBc) antigen resulted in nanoparticles displaying allergen-derived peptide upon its expression in prokaryotic host.• The resultant VLPs (chimeric HBc VLPs) did not exhibit IgE reactivity with allergic patients' sera and were not able to degranulate basophils.

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Constructing a hybrid molecule with low capacity of IgE binding from Chenopodium album pollen allergens

Cited by 14 publications

References 32 publications

Immunotherapy with a Recombinant Hybrid Molecule Alleviates Allergic Responses More Efficiently than an Allergenic Cocktail or Pollen Extract in a Model of <b><i>Chenopodium album</i></b> Allergy

Immunotherapy with a Recombinant Hybrid Molecule Alleviates Allergic Responses More Efficiently than an Allergenic Cocktail or Pollen Extract in a Model of <b><i>Chenopodium album</i></b> Allergy

Expression of a Chimeric Allergen with High Rare Codons Content in Codon Bias-Adjusted Escherichia coli: Escherichia coli BL21 (DE3)-Codon Plus RIL as an Efficient Host

Genetically engineered fusion of allergen and viral-like particle induces a more effective allergen-specific immune response than a combination of them

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