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dc.creatorApostolović, Danijela
dc.creatorLuykx, Dion
dc.creatorWarmenhoven, Hans
dc.creatorVerbart, Dennis
dc.creatorStanić-Vučinić, Dragana
dc.creatorde Jong, Govardus A. H.
dc.creatorĆirković-Veličković, Tanja
dc.creatorKoppelman, Stef J.
dc.date.accessioned2018-11-22T00:24:54Z
dc.date.available2018-11-22T00:24:54Z
dc.date.issued2013
dc.identifier.issn1570-9639
dc.identifier.urihttp://cherry.chem.bg.ac.rs/handle/123456789/1463
dc.description.abstractConglutins, the major peanut allergens, Ara h 2 and Ara h 6, are highly structured proteins stabilized by multiple disulfide bridges and are stable towards heat-denaturation and digestion. We sought a way to reduce their potent allergenicity in view of the development of immunotherapy for peanut allergy. Isoforms of conglutin were purified, reduced with dithiothreitol and subsequently allcylated with iodoacetamide. The effect of this modification was assessed on protein folding and IgE-binding. We found that all disulfide bridges were reduced and alkylated. As a result, the secondary structure lost a-helix and gained some beta-structure content, and the tertiary structure stability was reduced. On a functional level, the modification led to a strongly decreased IgE-binding. Using conditions for limited reduction and alkylation, partially reduced and alkylated proteins were found with rearranged disulfide bridges and, in some cases, intermolecular cross-links were found. Peptide mass finger printing was applied to control progress of the modification reaction and to map novel disulfide bonds. There was no preference for the order in which disulfides were reduced, and disulfide rearrangement occurred in a non-specific way. Only minor differences in kinetics of reduction and alkylation were found between the different conglutin isoforms. We conclude that the peanut conglutins Ara h 2 and Ara h 6 can be chemically modified by reduction and alkylation, such that they substantially unfold and that their allergenic potency decreases. (C) 2013 Elsevier B.V. All rights reserved.en
dc.publisherElsevier Science Bv, Amsterdam
dc.relationinfo:eu-repo/grantAgreement/MESTD/Basic Research (BR or ON)/172024/RS//
dc.relationinfo:eu-repo/grantAgreement/EC/FP7/256716/EU//
dc.rightsrestrictedAccess
dc.sourceBiochimica et Biophysica Acta: Proteins and Proteomics
dc.subjectAllergenen
dc.subjectMass spectrometry (MS)en
dc.subjectPeanuten
dc.subjectSpectroscopyen
dc.subjectImmunochemistryen
dc.subjectProtein structureen
dc.titleReduction and alkylation of peanut allergen isoforms Ara h 2 and Ara h 6 characterization of intermediate- and end productsen
dc.typearticle
dc.rights.licenseARR
dcterms.abstractКоппелман, Стеф Ј.; Вербарт, Деннис; Wарменховен, Ханс; Луyкx, Дион; Aпостоловиц, Данијела; Станић-Вучинић, Драгана; де Јонг, Говардус A. Х.; Ћирковић-Величковић, Тања;
dc.citation.volume1834
dc.citation.issue12
dc.citation.spage2832
dc.citation.epage2842
dc.identifier.wos000329418300040
dc.identifier.doi10.1016/j.bbapap.2013.10.004
dc.citation.other1834(12): 2832-2842
dc.citation.rankM22
dc.identifier.pmid24145103
dc.type.versionpublishedVersionen
dc.identifier.scopus2-s2.0-84887391859
dc.identifier.rcubKon_2583


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