Characterization of Human Plasma Glycoproteins Separated by Two-Dimensional Gel Electrophoresis
Purification of protein isoforms for the characterization of post-translational modifications, such as glycosylation, can be laborious and demanding. We report a means of determining monosaccharide composition and the identity of glycoproteins from a single spot on a two-dimensional (2-D) gel. The s...
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| Veröffentlicht in: | Bio/Technology 1996-01, Vol.14 (1), p.66-70 |
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| container_title | Bio/Technology |
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| creator | Packer, Nicolle H. Wilkins, Marc R. Golaz, Olivier Lawson, Margaret A. Gooley, Andrew A. Hochstrasser, Denis F. Redmond, John W. Williams, Keith L. |
| description | Purification of protein isoforms for the characterization of post-translational modifications, such as glycosylation, can be laborious and demanding. We report a means of determining monosaccharide composition and the identity of glycoproteins from a single spot on a two-dimensional (2-D) gel. The sensitivity of the method depends on the degree of glycosylation of the protein. We show that bovine fetuin can be analyzed and identified at the level of 100 pmol. 2-D reference maps enable quick identification of glycoprotein isoforms, and the nature of glycosylation differences. Human sera glycoforms were isolated by micropreparative 2-D PAGE using a narrow-range immobilized pH gradient. Single spots excised from one polyvinylidene difluoride blot of a 2-D gel were used sequentially for sialic acid analysis, neutral and amino sugar analysis, and finally amino acid analysis. The glycosylation variations in isoforms of human fetuin and α-1-antitrypsin were determined. The amino acid composition, in conjunction with protein pI and MW, successfully identified the glycoproteins. |
| doi_str_mv | 10.1038/nbt0196-66 |
| format | Article |
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We report a means of determining monosaccharide composition and the identity of glycoproteins from a single spot on a two-dimensional (2-D) gel. The sensitivity of the method depends on the degree of glycosylation of the protein. We show that bovine fetuin can be analyzed and identified at the level of 100 pmol. 2-D reference maps enable quick identification of glycoprotein isoforms, and the nature of glycosylation differences. Human sera glycoforms were isolated by micropreparative 2-D PAGE using a narrow-range immobilized pH gradient. Single spots excised from one polyvinylidene difluoride blot of a 2-D gel were used sequentially for sialic acid analysis, neutral and amino sugar analysis, and finally amino acid analysis. The glycosylation variations in isoforms of human fetuin and α-1-antitrypsin were determined. The amino acid composition, in conjunction with protein pI and MW, successfully identified the glycoproteins.</description><identifier>ISSN: 0733-222X</identifier><identifier>ISSN: 1087-0156</identifier><identifier>EISSN: 1546-1696</identifier><identifier>EISSN: 2331-3684</identifier><identifier>DOI: 10.1038/nbt0196-66</identifier><identifier>PMID: 9636314</identifier><language>eng</language><publisher>New York: Nature Publishing Group US</publisher><subject>Agriculture ; alpha 1-Antitrypsin - analysis ; alpha-Fetoproteins - analysis ; Analytical, structural and metabolic biochemistry ; Animals ; Bioinformatics ; Biological and medical sciences ; Biomedical and Life Sciences ; Biomedical Engineering/Biotechnology ; Biomedicine ; Biotechnology ; Cattle ; Electrophoresis, Gel, Two-Dimensional ; Fundamental and applied biological sciences. Psychology ; Glycoproteins ; Glycoproteins - blood ; Glycoproteins - isolation & purification ; Glycosylation ; Humans ; Life Sciences ; Monosaccharides - analysis ; Oligosaccharides - analysis ; Proteins</subject><ispartof>Bio/Technology, 1996-01, Vol.14 (1), p.66-70</ispartof><rights>Nature Publishing Company 1996</rights><rights>1996 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c414t-20888cb63eee17be5d57bf6099fcd558852a6113d0e5fd2fcf10fe283877e4073</citedby><cites>FETCH-LOGICAL-c414t-20888cb63eee17be5d57bf6099fcd558852a6113d0e5fd2fcf10fe283877e4073</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://link.springer.com/content/pdf/10.1038/nbt0196-66$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1038/nbt0196-66$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>314,776,780,4010,27900,27901,27902,41464,42533,51294</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=2981773$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/9636314$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Packer, Nicolle H.</creatorcontrib><creatorcontrib>Wilkins, Marc R.</creatorcontrib><creatorcontrib>Golaz, Olivier</creatorcontrib><creatorcontrib>Lawson, Margaret A.</creatorcontrib><creatorcontrib>Gooley, Andrew A.</creatorcontrib><creatorcontrib>Hochstrasser, Denis F.</creatorcontrib><creatorcontrib>Redmond, John W.</creatorcontrib><creatorcontrib>Williams, Keith L.</creatorcontrib><title>Characterization of Human Plasma Glycoproteins Separated by Two-Dimensional Gel Electrophoresis</title><title>Bio/Technology</title><addtitle>Nat Biotechnol</addtitle><addtitle>Biotechnology (N Y)</addtitle><description>Purification of protein isoforms for the characterization of post-translational modifications, such as glycosylation, can be laborious and demanding. We report a means of determining monosaccharide composition and the identity of glycoproteins from a single spot on a two-dimensional (2-D) gel. The sensitivity of the method depends on the degree of glycosylation of the protein. We show that bovine fetuin can be analyzed and identified at the level of 100 pmol. 2-D reference maps enable quick identification of glycoprotein isoforms, and the nature of glycosylation differences. Human sera glycoforms were isolated by micropreparative 2-D PAGE using a narrow-range immobilized pH gradient. Single spots excised from one polyvinylidene difluoride blot of a 2-D gel were used sequentially for sialic acid analysis, neutral and amino sugar analysis, and finally amino acid analysis. The glycosylation variations in isoforms of human fetuin and α-1-antitrypsin were determined. The amino acid composition, in conjunction with protein pI and MW, successfully identified the glycoproteins.</description><subject>Agriculture</subject><subject>alpha 1-Antitrypsin - analysis</subject><subject>alpha-Fetoproteins - analysis</subject><subject>Analytical, structural and metabolic biochemistry</subject><subject>Animals</subject><subject>Bioinformatics</subject><subject>Biological and medical sciences</subject><subject>Biomedical and Life Sciences</subject><subject>Biomedical Engineering/Biotechnology</subject><subject>Biomedicine</subject><subject>Biotechnology</subject><subject>Cattle</subject><subject>Electrophoresis, Gel, Two-Dimensional</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Glycoproteins</subject><subject>Glycoproteins - blood</subject><subject>Glycoproteins - isolation & purification</subject><subject>Glycosylation</subject><subject>Humans</subject><subject>Life Sciences</subject><subject>Monosaccharides - analysis</subject><subject>Oligosaccharides - analysis</subject><subject>Proteins</subject><issn>0733-222X</issn><issn>1087-0156</issn><issn>1546-1696</issn><issn>2331-3684</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1996</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkUFv1DAQhS0EKtuFC3ckHxAHUMB2HNs5Vku7RaoEEkXiZjnOmKZy4sWTCC2_Hlcb9YTEaQ7vmzczbwh5xdkHzmrzcepmxltVKfWEbHgjVcVVq56SDdN1XQkhfjwn54j3jEmthDwjZ62qVc3lhtjdncvOz5CHP24e0kRToNfL6Cb6NTocHd3Ho0-HnGYYJqTf4FD4GXraHent71R9GkaYsDS6SPcQ6WUEP-d0uEsZcMAX5FlwEeHlWrfk-9Xl7e66uvmy_7y7uKm85HKuBDPG-E7VAMB1B03f6C4o1rbB901jTCOc4rzuGTShF8EHzgIIUxutQZYzt-Ttybds-msBnO04oIcY3QRpQauNVkYy81-Ql2lSqQfHdyfQ54SYIdhDHkaXj5Yz-xC7XWO3ShX49eq6dCP0j-iac9HfrLpD72LIbvIDPmKiNVyXX23J-xOGRZl-Qrb3acklWvzX0L_Ks5my</recordid><startdate>19960101</startdate><enddate>19960101</enddate><creator>Packer, Nicolle H.</creator><creator>Wilkins, Marc R.</creator><creator>Golaz, Olivier</creator><creator>Lawson, Margaret A.</creator><creator>Gooley, Andrew A.</creator><creator>Hochstrasser, Denis F.</creator><creator>Redmond, John W.</creator><creator>Williams, Keith L.</creator><general>Nature Publishing Group US</general><general>Nature Publications</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QO</scope><scope>8FD</scope><scope>FR3</scope><scope>P64</scope><scope>7X8</scope></search><sort><creationdate>19960101</creationdate><title>Characterization of Human Plasma Glycoproteins Separated by Two-Dimensional Gel Electrophoresis</title><author>Packer, Nicolle H. ; Wilkins, Marc R. ; Golaz, Olivier ; Lawson, Margaret A. ; Gooley, Andrew A. ; Hochstrasser, Denis F. ; Redmond, John W. ; Williams, Keith L.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c414t-20888cb63eee17be5d57bf6099fcd558852a6113d0e5fd2fcf10fe283877e4073</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1996</creationdate><topic>Agriculture</topic><topic>alpha 1-Antitrypsin - analysis</topic><topic>alpha-Fetoproteins - analysis</topic><topic>Analytical, structural and metabolic biochemistry</topic><topic>Animals</topic><topic>Bioinformatics</topic><topic>Biological and medical sciences</topic><topic>Biomedical and Life Sciences</topic><topic>Biomedical Engineering/Biotechnology</topic><topic>Biomedicine</topic><topic>Biotechnology</topic><topic>Cattle</topic><topic>Electrophoresis, Gel, Two-Dimensional</topic><topic>Fundamental and applied biological sciences. 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We report a means of determining monosaccharide composition and the identity of glycoproteins from a single spot on a two-dimensional (2-D) gel. The sensitivity of the method depends on the degree of glycosylation of the protein. We show that bovine fetuin can be analyzed and identified at the level of 100 pmol. 2-D reference maps enable quick identification of glycoprotein isoforms, and the nature of glycosylation differences. Human sera glycoforms were isolated by micropreparative 2-D PAGE using a narrow-range immobilized pH gradient. Single spots excised from one polyvinylidene difluoride blot of a 2-D gel were used sequentially for sialic acid analysis, neutral and amino sugar analysis, and finally amino acid analysis. The glycosylation variations in isoforms of human fetuin and α-1-antitrypsin were determined. 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| identifier | ISSN: 0733-222X |
| ispartof | Bio/Technology, 1996-01, Vol.14 (1), p.66-70 |
| issn | 0733-222X 1087-0156 1546-1696 2331-3684 |
| language | eng |
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| source | MEDLINE; Nature Journals Online; SpringerLink Journals - AutoHoldings |
| subjects | Agriculture alpha 1-Antitrypsin - analysis alpha-Fetoproteins - analysis Analytical, structural and metabolic biochemistry Animals Bioinformatics Biological and medical sciences Biomedical and Life Sciences Biomedical Engineering/Biotechnology Biomedicine Biotechnology Cattle Electrophoresis, Gel, Two-Dimensional Fundamental and applied biological sciences. Psychology Glycoproteins Glycoproteins - blood Glycoproteins - isolation & purification Glycosylation Humans Life Sciences Monosaccharides - analysis Oligosaccharides - analysis Proteins |
| title | Characterization of Human Plasma Glycoproteins Separated by Two-Dimensional Gel Electrophoresis |
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