Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment
Blood plasma is considered to be an excellent source of disease biomarkers because it contains proteins, lipids, metabolites, cell, and cell-derived extracellular vesicles from different cellular origins including diseased tissues. Most secretory and membranous proteins that can be found in plasma a...
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| Format: | Journal Article |
| Language: | English |
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2016
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| Online Access: | https://hdl.handle.net/10356/81887 http://hdl.handle.net/10220/39735 |
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| author | Adav, Sunil S. Hwa, Ho Hee de Kleijn, Dominique Sze, Siu Kwan |
| author2 | School of Biological Sciences |
| author_facet | School of Biological Sciences Adav, Sunil S. Hwa, Ho Hee de Kleijn, Dominique Sze, Siu Kwan |
| author_sort | Adav, Sunil S. |
| collection | NTU |
| description | Blood plasma is considered to be an excellent source of disease biomarkers because it contains proteins, lipids, metabolites, cell, and cell-derived extracellular vesicles from different cellular origins including diseased tissues. Most secretory and membranous proteins that can be found in plasma are glycoproteins; therefore, the plasma glycoproteome is one of the major subproteomes that is highly enriched with disease biomarkers. As a result, the glycoproteome has attracted much attention in clinical proteomic research. The modification of proteins with glycans regulates a wide range of functions in biology, but profiling plasma glycoproteins on a global scale has been hampered by the presence of low stoichiometry of glycoproteins in a complex high abundance plasma proteome background and lack of effective analytical technique. This study aims to improve plasma glycoproteome coverage using pig plasma as a model sample with a two-step strategy. The first step involves fractionation of the plasma proteins using ultracentrifugation into supernatant and pellet that is believed to contain low abundant glycoproteins. In the second step, further enrichment of glycopeptides was achieved in both fractions by adopting electrostatic repulsion hydrophilic interaction chromatography (ERLIC) coupled to tandem mass spectrometry (LC-MS/MS) analysis. The coverage of enriched glycoproteins in supernatant, pellet, and whole plasma sample as control was compared. Using this simple sample fractionation approach by ultracentrifugation and further ERLIC enrichment technique, sample complexity was reduced and glycoproteome coverage was significantly enhanced in supernatant and pellet fractions (by >50%) compared with whole plasma sample. This study showed that when ultracentrifugation is coupled to ERLIC glycopeptides enrichment and glycoproteome identification are significantly improved. This study demonstrates the combination of ultracentrifugation and ERLIC as a useful method for discovering plasma glycoprotein disease biomarkers. |
| first_indexed | 2024-10-01T06:39:27Z |
| format | Journal Article |
| id | ntu-10356/81887 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T06:39:27Z |
| publishDate | 2016 |
| record_format | dspace |
| spelling | ntu-10356/818872023-02-28T16:59:11Z Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment Adav, Sunil S. Hwa, Ho Hee de Kleijn, Dominique Sze, Siu Kwan School of Biological Sciences ERLIC; glycoproteins; plasma proteome; protein glycosylation; biomarkers Blood plasma is considered to be an excellent source of disease biomarkers because it contains proteins, lipids, metabolites, cell, and cell-derived extracellular vesicles from different cellular origins including diseased tissues. Most secretory and membranous proteins that can be found in plasma are glycoproteins; therefore, the plasma glycoproteome is one of the major subproteomes that is highly enriched with disease biomarkers. As a result, the glycoproteome has attracted much attention in clinical proteomic research. The modification of proteins with glycans regulates a wide range of functions in biology, but profiling plasma glycoproteins on a global scale has been hampered by the presence of low stoichiometry of glycoproteins in a complex high abundance plasma proteome background and lack of effective analytical technique. This study aims to improve plasma glycoproteome coverage using pig plasma as a model sample with a two-step strategy. The first step involves fractionation of the plasma proteins using ultracentrifugation into supernatant and pellet that is believed to contain low abundant glycoproteins. In the second step, further enrichment of glycopeptides was achieved in both fractions by adopting electrostatic repulsion hydrophilic interaction chromatography (ERLIC) coupled to tandem mass spectrometry (LC-MS/MS) analysis. The coverage of enriched glycoproteins in supernatant, pellet, and whole plasma sample as control was compared. Using this simple sample fractionation approach by ultracentrifugation and further ERLIC enrichment technique, sample complexity was reduced and glycoproteome coverage was significantly enhanced in supernatant and pellet fractions (by >50%) compared with whole plasma sample. This study showed that when ultracentrifugation is coupled to ERLIC glycopeptides enrichment and glycoproteome identification are significantly improved. This study demonstrates the combination of ultracentrifugation and ERLIC as a useful method for discovering plasma glycoprotein disease biomarkers. MOE (Min. of Education, S’pore) Accepted version 2016-01-21T03:47:13Z 2019-12-06T14:42:23Z 2016-01-21T03:47:13Z 2019-12-06T14:42:23Z 2015 Journal Article Adav, S. S., Hwa, H. H., de Kleijn, D.,& Sze, S. K. (2015). Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment. Journal of Proteome Research, 14(7), 2828-2838. Adav, S. S., Hwa, H. H., de Kleijn, D., & Sze, S. K. (2015). Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment. Journal of Proteome Research, 14(7), 2828-2838. 1535-3893 https://hdl.handle.net/10356/81887 http://hdl.handle.net/10220/39735 10.1021/acs.jproteome.5b00102 en Journal of Proteome Research © 2015 American Chemical Society. This is the author created version of a work that has been peer reviewed and accepted for publication by Journal of Proteome Research, American Chemical Society. It incorporates referee’s comments but changes resulting from the publishing process, such as copyediting, structural formatting, may not be reflected in this document. The published version is available at: [http://dx.doi.org/10.1021/acs.jproteome.5b00102]. 30 p. application/pdf |
| spellingShingle | ERLIC; glycoproteins; plasma proteome; protein glycosylation; biomarkers Adav, Sunil S. Hwa, Ho Hee de Kleijn, Dominique Sze, Siu Kwan Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment |
| title | Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment |
| title_full | Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment |
| title_fullStr | Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment |
| title_full_unstemmed | Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment |
| title_short | Improving Blood Plasma Glycoproteome Coverage by Coupling Ultracentrifugation Fractionation to Electrostatic Repulsion–Hydrophilic Interaction Chromatography Enrichment |
| title_sort | improving blood plasma glycoproteome coverage by coupling ultracentrifugation fractionation to electrostatic repulsion hydrophilic interaction chromatography enrichment |
| topic | ERLIC; glycoproteins; plasma proteome; protein glycosylation; biomarkers |
| url | https://hdl.handle.net/10356/81887 http://hdl.handle.net/10220/39735 |
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