Evaluation of the Allergenicity Potential of TcPR-10 Protein from Theobroma cacao 英文参考文献.docVIP

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Evaluation of the Allergenicity Potential of TcPR-10 Protein from Theobroma cacao 英文参考文献.doc

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Evaluation of the Allergenicity Potential of TcPR-10 Protein from Theobroma cacao 英文参考文献

EvaluationoftheAllergenicityPotentialofTcPR-10 ProteinfromTheobromacacao SaraPereiraMenezes1.,JaneLimadosSantos2.,ThyagoHermyllySantanaCardoso1 ,Carlos PriminhoPirovani1,FabienneMicheli1,3,Fa′timaSoaresMottaNoronha4,Andre′aCata?oAlves4 ,Ana MariaCaetanoFaria4,AbelmondaSilvaGesteira5* 1UESC,CentrodeBiotecnologiaeGene′tica,Ilhe′us,Bahia,Brasil,2UESC,CentrodeMicroscopiaEletro?nica,Ilhe′us,Bahia,Brasil,3CIRAD,UMARAGAP,Montpellier,France, 4UFMG,InstitutodeCie?ncias Biolo′gicas,Pampulha,BeloHorizonte,MinasGerais,Brasil,5EmbrapaMandiocaeFruticultura,CruzdasAlmas,Bahia,Brasil Abstract Background: The pathogenesis related protein PR10 (TcPR-10), obtained from the Theobroma cacao-Moniliophthora perniciosainteractionlibrary,presentsantifungalactivityagainstM.perniciosaandactsinvitroasaribonuclease.However, despiteitsbiotechnologicalpotential,theTcPR-10hastheP-loopmotifsimilartothoseofsomeallergenicproteinssuchas Betv1(Betulaverrucosa)andPruav1(Prunusavium).Theinsertionofmutationsinthismotifcanproduceproteinswith reducedallergenicpower.Theobjectiveofthepresentworkwastoevaluatetheallergenicpotentialofthewildtypeand mutantrecombinantTcPR-10usingbioinformaticstoolsandimmunologicalassays. Methodology/Principal Findings: Mutant substitutions (T10P, I30V, H45S) were inserted in the TcPR-10 gene by site- directedmutagenesis,clonedintopET28aandexpressedinEscherichiacoliBL21(DE3)cells.Changesinmolecularsurface causedbythemutantsubstitutionswasevaluatedbycomparativeproteinmodelingusingthethree-dimensionalstructure ofthemajorcherryallergen,Pruav1asatemplate.Theimmunologicalassayswerecarriedoutin8–12weekoldfemale BALB/c mice. The mice were sensitized with the proteins (wild type and mutants) via subcutaneous and challenged intranasalforinductionofallergicairwayinflammation. Conclusions/Significance: We showed that the wild TcPR-10 protein has allergenic potential, whereas the insertion of mutationsproducedproteinswithreducedcapacityofIgEproductionandcellularinfiltrationinthelungs.Onthe