Quantification, 2de analysis and identification of enriched glycosylated proteins from mouse muscles: difficulties and alternatives

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dc.contributor.authorMenegoci Eugenio, Patricia de Fatima
dc.contributor.authorAssunção, Nilson Antonio [UNIFESP]
dc.contributor.authorSciandra, Francesca
dc.contributor.authorAquino, Adriano
dc.contributor.authorBrancaccio, Andrea
dc.contributor.authorCarrilho, Emanuel
dc.date.accessioned2019-01-21T10:30:15Z
dc.date.available2019-01-21T10:30:15Z
dc.date.issued2016
dc.description.abstractOne of the problems with 2DE is that proteins present in low amounts in a sample are usually not detected, since their signals are masked by the predominant proteins. The elimination of these abundant proteins is not a guaranteed solution to achieve the desired results. The main objective of this study was the comparison of common and simple methodologies employed for 2DE analysis followed by MS identification, focusing on a pre-purified sample using a wheat germ agglutinin (WGA) column. Adult male C57Black/Crj6 (C57BL/6) mice were chosen as the model animal in this studyen
dc.description.abstractthe gastrocnemius muscles were collected and processed for the experiments. The initial fractionation with succinylated WGA was successful for the elimination of the most abundant proteins. Two quantification methods were employed for the purified samples, and bicinchoninic acid (BCA) was proven to be most reliable for the quantification of glycoproteins. The gel staining method, however, was found to be decisive for the detection of specific proteins, since their structures affect the interaction of the dye with the peptide backbone. The Coomassie Blue R-250 dye very weakly stained the gel with the WGA purified sample. When the same gel was stained with silver nitrate, however, MS could positively assign 12 new spots. The structure of the referred proteins was not found to be prone to interaction with Coomassie blue.en
dc.description.affiliationInstituto de Química de São Carlos, Universidade de São Paulo, São Carlos, SP, Brazil
dc.description.affiliationInstituto Nacional de Ciência e Tecnologia de Bioanalítica, Campinas, SP, Brazil
dc.description.affiliationInstituto de Ciências Ambientais, Químicas e Farmacêuticas, Universidade Federal de São Paulo, Diadema, SP, Brazil
dc.description.affiliationIstituto di Chimica del Riconoscimento Molecolare (CNR), c/o Istituto di Biochimica e Biochimica Clinica, Università Cattolica del Sacro Cuore, Roma, Italy
dc.description.affiliationUnifespInstituto de Ciências Ambientais, Químicas e Farmacêuticas, Universidade Federal de São Paulo, Diadema, SP, Brazil
dc.description.sourceWeb of Science
dc.description.sponsorshipFAPESP [2008/04050-4, 2009/54040-8, 2009/16598-7]
dc.description.sponsorshipCNPq [140840/2008-7, 160528/2011-9]
dc.description.sponsorshipCAPES [0203-10-6]
dc.description.sponsorshipIDFAPESP: 2008/04050-4
dc.description.sponsorshipIDFAPESP: 2009/54040-8
dc.description.sponsorshipIDFAPESP: 2009/16598-7
dc.description.sponsorshipIDCNPq: 140840/2008-7
dc.description.sponsorshipIDCNPq: 160528/2011-9
dc.description.sponsorshipIDCAPES: 0203-10-6
dc.format.extent321-334
dc.identifierhttps://doi.org/10.1002/elps.201500362
dc.identifier.citationElectrophoresis. Hoboken, v. 37, n. 2, p. 321-334, 2016.
dc.identifier.doi10.1002/elps.201500362
dc.identifier.issn0173-0835
dc.identifier.urihttp://repositorio.unifesp.br/handle/11600/49657
dc.identifier.wosWOS:000368030000011
dc.language.isoeng
dc.publisherWiley-blackwell
dc.relation.ispartofElectrophoresis
dc.rightsinfo:eu-repo/semantics/restrictedAccess
dc.subjectGel-Staining Methodsen
dc.subjectGlycoproteinen
dc.subjectProtein Quantificationen
dc.subjectTwo-Dimensional Gel ElectrophoresisLectin Affinity-Chromatographyen
dc.subject2-Dimensional Gel-Electrophoresisen
dc.subjectCoomassie Brilliant Blueen
dc.subjectWheat-Germ-Agglutininen
dc.subjectMass-Spectrometryen
dc.subjectMembrane-Glycoproteinsen
dc.subjectMuscular-Dystrophiesen
dc.subjectBiomarker Discoveryen
dc.subjectPolyacrylamide-Gelsen
dc.subjectBicinchoninic Aciden
dc.titleQuantification, 2de analysis and identification of enriched glycosylated proteins from mouse muscles: difficulties and alternativesen
dc.typeinfo:eu-repo/semantics/article
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