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dc.contributor.authorLin, Chi-Hung
dc.contributor.authorChik, Jenny HL
dc.contributor.authorPacker, Nicolle H
dc.contributor.authorMolloy, Mark P
dc.date.accessioned2017-05-11T04:22:13Z
dc.date.available2017-05-11T04:22:13Z
dc.date.issued2015
dc.identifier.issn1535-3893
dc.identifier.doi10.1021/pr500785f
dc.identifier.urihttp://hdl.handle.net/10072/336545
dc.description.abstractGlycosylation results from the concerted action of glycosylation enzymes in the secretory pathway. In general, gene expression serves as the primary control mechanism, but post-translational fine-tuning of glycosylation enzyme functions is often necessary for efficient synthesis of specific glycan epitopes. While the field of glycomics has rapidly advanced, there lacks routine proteomic methods to measure expression of specific glycosylation enzymes needed to fill the gap between mRNA expression and the glycomic profile in a “reverse genomics” workflow. Toward developing this workflow we enriched Golgi membranes from two human colon cancer cell lines by sucrose density centrifugation and further mass-based fractionation by SDS-PAGE. We then applied mass spectrometry to demonstrate a doubling in the number of Golgi resident proteins identified, compared to the unenriched, low speed centrifuged supernatant of lysed cells. A total of 35 Golgi-resident glycosylation enzymes, of which 23 were glycosyltransferases, were identified making this the largest protein database so far of Golgi resident glycosylation enzymes experimentally identified in cultured human cells. We developed targeted mass spectrometry assays for specific quantitation of many of these glycosylation enzymes. Our results show that alterations in abundance of glycosylation enzymes at the protein level were generally consistent with the resultant glycomic profiles, but not necessarily with the corresponding glycosyltransferase mRNA expression as exemplified by the case of O-glycan core 1 T synthase.
dc.description.peerreviewedYes
dc.languageEnglish
dc.language.isoeng
dc.publisherAmerican Chemical Society
dc.relation.ispartofpagefrom747
dc.relation.ispartofpageto755
dc.relation.ispartofissue2
dc.relation.ispartofjournalJournal of Proteome Research
dc.relation.ispartofvolume14
dc.subject.fieldofresearchBiochemistry and Cell Biology not elsewhere classified
dc.subject.fieldofresearchChemical Sciences
dc.subject.fieldofresearchBiological Sciences
dc.subject.fieldofresearchcode060199
dc.subject.fieldofresearchcode03
dc.subject.fieldofresearchcode06
dc.titleMultidimensional fractionation is a requirement for quantitation of golgi-resident glycosylation enzymes from cultured human cells
dc.typeJournal article
dc.type.descriptionC1 - Articles
dc.type.codeC - Journal Articles
gro.hasfulltextNo Full Text
gro.griffith.authorPacker, Nicki
gro.griffith.authorLin, Chi-Hung


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