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Native Separation and Metallation Analysis of SOD1 Protein from the Human Central Nervous System: a Methodological Workflow

hal.structure.identifierCentre d'Etudes Nucléaires de Bordeaux Gradignan [CENBG]
dc.contributor.authorROUDEAU, Stéphane
hal.structure.identifierThe University of Sydney
dc.contributor.authorTRIST, Benjamin G.
hal.structure.identifierCentre d'Etudes Nucléaires de Bordeaux Gradignan [CENBG]
dc.contributor.authorCARMONA, Asuncion
hal.structure.identifierThe University of Sydney
dc.contributor.authorDAVIES, Katherine M.
hal.structure.identifierThe University of Sydney
dc.contributor.authorHALLIDAY, Glenda M.
hal.structure.identifierUniversité de Bordeaux [UB]
dc.contributor.authorRUFIN, Yann
hal.structure.identifierUniversité de Bordeaux [UB]
hal.structure.identifierPlateforme Proteome, Univ. Bordeaux, F- 33076 Bordeaux, France
dc.contributor.authorCLAVEROL, Stéphane
hal.structure.identifierDeutsches Elektronen-Synchrotron [Hamburg] [DESY]
dc.contributor.authorVAN MALDEREN, Stijn J.M.
hal.structure.identifierDeutsches Elektronen-Synchrotron [Hamburg] [DESY]
dc.contributor.authorFALKENBERG, Gerald
hal.structure.identifierThe University of Sydney
dc.contributor.authorDOUBLE, Kay L.
hal.structure.identifierCentre d'Etudes Nucléaires de Bordeaux Gradignan [CENBG]
dc.contributor.authorORTEGA, Richard
dc.date.issued2021-08-04
dc.identifier.issn0003-2700
dc.description.abstractEnStudies of the metal content of metalloproteins in tissues from the human central nervous system (CNS) can be compromised by preparative techniques which alter levels of, or interactions between, metals and the protein of interest within a complex mixture. We developed a methodological workflow combining size exclusion chromatography, native isoelectric focusing, and either proton or synchrotron X-ray fluorescence within electrophoresis gels to analyze the endogenous metal content of copper-zinc superoxide dismutase (SOD1) purified from minimal amounts (<20 mg) of post-mortem human brain and spinal cord tissue. Abnormal metallation and aggregation of SOD1 are suspected to play a role in amyotrophic lateral sclerosis and Parkinson’s disease, but data describing SOD1 metal occupancy in human tissues have not previously been reported. Validating our novel approach, we demonstrated step-by-step metal preservation, preserved SOD1 activity, and substantial enrichment of SOD1 protein versus confounding metalloproteins. We analyzed tissues from nine healthy individuals and five CNS regions (occipital cortex, substantia nigra, locus coeruleus, dorsal spinal cord, and ventral spinal cord). We found that Cu and Zn were bound to SOD1 in a ratio of 1.12 ± 0.28, a ratio very close to the expected value of 1. Our methodological workflow can be applied to the study of endogenous native SOD1 in a pathological context and adapted to a range of metalloproteins from human tissues and other sources.
dc.language.isoen
dc.publisherAmerican Chemical Society
dc.subject.enSuperoxide dismutase 1 SOD1
dc.subject.encopper
dc.subject.enzinc
dc.subject.ensynchrotron X-ray fluorescence
dc.subject.encentral nervous system
dc.title.enNative Separation and Metallation Analysis of SOD1 Protein from the Human Central Nervous System: a Methodological Workflow
dc.typeArticle de revue
dc.identifier.doi10.1021/acs.analchem.1c01128
dc.subject.halChimie/Chimie analytique
dc.subject.halSciences du Vivant [q-bio]/Biochimie, Biologie Moléculaire/Biochimie [q-bio.BM]
bordeaux.journalAnalytical Chemistry
bordeaux.page11108-11115
bordeaux.volume93
bordeaux.issue32
bordeaux.peerReviewedoui
hal.identifierhal-03326181
hal.version1
hal.popularnon
hal.audienceInternationale
hal.origin.linkhttps://hal.archives-ouvertes.fr//hal-03326181v1
bordeaux.COinSctx_ver=Z39.88-2004&amp;rft_val_fmt=info:ofi/fmt:kev:mtx:journal&amp;rft.jtitle=Analytical%20Chemistry&amp;rft.date=2021-08-04&amp;rft.volume=93&amp;rft.issue=32&amp;rft.spage=11108-11115&amp;rft.epage=11108-11115&amp;rft.eissn=0003-2700&amp;rft.issn=0003-2700&amp;rft.au=ROUDEAU,%20St%C3%A9phane&amp;TRIST,%20Benjamin%20G.&amp;CARMONA,%20Asuncion&amp;DAVIES,%20Katherine%20M.&amp;HALLIDAY,%20Glenda%20M.&amp;rft.genre=article


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