Simultaneous determination of nucleotide sugars with ion-pair reversed-phase HPLC
Nucleotide sugars are important in determining cell surface glycoprotein glycosylation, which can modulate cellular properties such as growth and arrest. We have developed a conventional HPLC method for simultaneous determination of nucleotide sugars. A mixture of nucleotide sugars (CMP-NeuAc, UDP-G...
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| Veröffentlicht in: | Glycobiology (Oxford) 2010-07, Vol.20 (7), p.865-871 |
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| container_title | Glycobiology (Oxford) |
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| creator | Nakajima, Kazuki Kitazume, Shinobu Angata, Takashi Fujinawa, Reiko Ohtsubo, Kazuaki Miyoshi, Eiji Taniguchi, Naoyuki |
| description | Nucleotide sugars are important in determining cell surface glycoprotein glycosylation, which can modulate cellular properties such as growth and arrest. We have developed a conventional HPLC method for simultaneous determination of nucleotide sugars. A mixture of nucleotide sugars (CMP-NeuAc, UDP-Gal, UDP-Glc, UDP-GalNAc, UDP-GlcNAc, GDP-Man, GDP-Fuc and UDP-GlcUA) and relevant nucleotides were perfectly separated in an optimized ion-pair reversed-phase mode using Inertsil ODS-4 and ODS-3 columns. The newly developed method enabled us to determine the nucleotide sugars in cellular extracts from 1 x 10⁶ cells in a single run. We applied this method to characterize nucleotide sugar levels in breast and pancreatic cancer cell lines and revealed that the abundance of UDP-GlcNAc, UDP-GalNAc, UDP-GlcUA and GDP-Fuc were a cell-type-specific feature. To determine the physiological significance of changes in nucleotide sugar levels, we analyzed their changes by glucose deprivation and found that the determination of nucleotide sugar levels provided us with valuable information with respect to studying the overview of cellular glycosylation status. |
| doi_str_mv | 10.1093/glycob/cwq044 |
| format | Article |
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We have developed a conventional HPLC method for simultaneous determination of nucleotide sugars. A mixture of nucleotide sugars (CMP-NeuAc, UDP-Gal, UDP-Glc, UDP-GalNAc, UDP-GlcNAc, GDP-Man, GDP-Fuc and UDP-GlcUA) and relevant nucleotides were perfectly separated in an optimized ion-pair reversed-phase mode using Inertsil ODS-4 and ODS-3 columns. The newly developed method enabled us to determine the nucleotide sugars in cellular extracts from 1 x 10⁶ cells in a single run. We applied this method to characterize nucleotide sugar levels in breast and pancreatic cancer cell lines and revealed that the abundance of UDP-GlcNAc, UDP-GalNAc, UDP-GlcUA and GDP-Fuc were a cell-type-specific feature. To determine the physiological significance of changes in nucleotide sugar levels, we analyzed their changes by glucose deprivation and found that the determination of nucleotide sugar levels provided us with valuable information with respect to studying the overview of cellular glycosylation status.</description><identifier>ISSN: 0959-6658</identifier><identifier>EISSN: 1460-2423</identifier><identifier>DOI: 10.1093/glycob/cwq044</identifier><identifier>PMID: 20371511</identifier><language>eng</language><publisher>England: Oxford University Press</publisher><subject>Carbohydrates - chemistry ; Cell Line, Tumor ; Chromatography, High Pressure Liquid - methods ; Chromatography, Ion Exchange - methods ; Glycosylation ; Humans ; ion-pair RP-HPLC ; Membrane Glycoproteins - chemistry ; Membrane Glycoproteins - metabolism ; nucleotide sugar ; Nucleotides - chemistry ; Nucleotides - metabolism ; Reproducibility of Results ; Sensitivity and Specificity</subject><ispartof>Glycobiology (Oxford), 2010-07, Vol.20 (7), p.865-871</ispartof><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c425t-3c7e46fe0313b362ccc90a33d8de4ae3d68db85c370f2b47a0e50bb24a3492423</citedby><cites>FETCH-LOGICAL-c425t-3c7e46fe0313b362ccc90a33d8de4ae3d68db85c370f2b47a0e50bb24a3492423</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27922,27923</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/20371511$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Nakajima, Kazuki</creatorcontrib><creatorcontrib>Kitazume, Shinobu</creatorcontrib><creatorcontrib>Angata, Takashi</creatorcontrib><creatorcontrib>Fujinawa, Reiko</creatorcontrib><creatorcontrib>Ohtsubo, Kazuaki</creatorcontrib><creatorcontrib>Miyoshi, Eiji</creatorcontrib><creatorcontrib>Taniguchi, Naoyuki</creatorcontrib><title>Simultaneous determination of nucleotide sugars with ion-pair reversed-phase HPLC</title><title>Glycobiology (Oxford)</title><addtitle>Glycobiology</addtitle><description>Nucleotide sugars are important in determining cell surface glycoprotein glycosylation, which can modulate cellular properties such as growth and arrest. We have developed a conventional HPLC method for simultaneous determination of nucleotide sugars. A mixture of nucleotide sugars (CMP-NeuAc, UDP-Gal, UDP-Glc, UDP-GalNAc, UDP-GlcNAc, GDP-Man, GDP-Fuc and UDP-GlcUA) and relevant nucleotides were perfectly separated in an optimized ion-pair reversed-phase mode using Inertsil ODS-4 and ODS-3 columns. The newly developed method enabled us to determine the nucleotide sugars in cellular extracts from 1 x 10⁶ cells in a single run. We applied this method to characterize nucleotide sugar levels in breast and pancreatic cancer cell lines and revealed that the abundance of UDP-GlcNAc, UDP-GalNAc, UDP-GlcUA and GDP-Fuc were a cell-type-specific feature. To determine the physiological significance of changes in nucleotide sugar levels, we analyzed their changes by glucose deprivation and found that the determination of nucleotide sugar levels provided us with valuable information with respect to studying the overview of cellular glycosylation status.</description><subject>Carbohydrates - chemistry</subject><subject>Cell Line, Tumor</subject><subject>Chromatography, High Pressure Liquid - methods</subject><subject>Chromatography, Ion Exchange - methods</subject><subject>Glycosylation</subject><subject>Humans</subject><subject>ion-pair RP-HPLC</subject><subject>Membrane Glycoproteins - chemistry</subject><subject>Membrane Glycoproteins - metabolism</subject><subject>nucleotide sugar</subject><subject>Nucleotides - chemistry</subject><subject>Nucleotides - metabolism</subject><subject>Reproducibility of Results</subject><subject>Sensitivity and Specificity</subject><issn>0959-6658</issn><issn>1460-2423</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2010</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkU1P3DAQhq2qqCy0x17b3HoKjD22kxzpUlikRbQCpKoXy3Emi2myWeyEj39PUPg4cprDPHo17zOMfeWwx6HA_VXz4Lpy393dgJQf2IxLDamQAj-yGRSqSLVW-TbbifEagGueq09sWwBmXHE-Y3_OfTs0vV1TN8Skop5C69e299066epkPbiGut5XlMRhZUNM7nx_lYzbdGN9SALdUohUpZsrGylZ_F7OP7Ot2jaRvjzPXXZ59OtivkiXZ8cn84Nl6qRQfYouI6lrAuRYohbOuQIsYpVXJC1hpfOqzJXDDGpRyswCKShLIS3K4qnfLvsx5W5CdzNQ7E3ro6OmmbqYTGrkY1F8n8TxhgxBjWQ6kS50MQaqzSb41oYHw8E86TaTbjPpHvlvz8lD2VL1Sr_4fQv0saf7170N_43OMFNm8fef4fnphfjJD8185L9PfG07Y1fBR3N5LoAjjI-DXAI-Asp9lbE</recordid><startdate>20100701</startdate><enddate>20100701</enddate><creator>Nakajima, Kazuki</creator><creator>Kitazume, Shinobu</creator><creator>Angata, Takashi</creator><creator>Fujinawa, Reiko</creator><creator>Ohtsubo, Kazuaki</creator><creator>Miyoshi, Eiji</creator><creator>Taniguchi, Naoyuki</creator><general>Oxford University Press</general><scope>FBQ</scope><scope>BSCLL</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>7X8</scope><scope>7TM</scope></search><sort><creationdate>20100701</creationdate><title>Simultaneous determination of nucleotide sugars with ion-pair reversed-phase HPLC</title><author>Nakajima, Kazuki ; Kitazume, Shinobu ; Angata, Takashi ; Fujinawa, Reiko ; Ohtsubo, Kazuaki ; Miyoshi, Eiji ; Taniguchi, Naoyuki</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c425t-3c7e46fe0313b362ccc90a33d8de4ae3d68db85c370f2b47a0e50bb24a3492423</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2010</creationdate><topic>Carbohydrates - chemistry</topic><topic>Cell Line, Tumor</topic><topic>Chromatography, High Pressure Liquid - methods</topic><topic>Chromatography, Ion Exchange - methods</topic><topic>Glycosylation</topic><topic>Humans</topic><topic>ion-pair RP-HPLC</topic><topic>Membrane Glycoproteins - chemistry</topic><topic>Membrane Glycoproteins - metabolism</topic><topic>nucleotide sugar</topic><topic>Nucleotides - chemistry</topic><topic>Nucleotides - metabolism</topic><topic>Reproducibility of Results</topic><topic>Sensitivity and Specificity</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Nakajima, Kazuki</creatorcontrib><creatorcontrib>Kitazume, Shinobu</creatorcontrib><creatorcontrib>Angata, Takashi</creatorcontrib><creatorcontrib>Fujinawa, Reiko</creatorcontrib><creatorcontrib>Ohtsubo, Kazuaki</creatorcontrib><creatorcontrib>Miyoshi, Eiji</creatorcontrib><creatorcontrib>Taniguchi, Naoyuki</creatorcontrib><collection>AGRIS</collection><collection>Istex</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>Nucleic Acids Abstracts</collection><jtitle>Glycobiology (Oxford)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Nakajima, Kazuki</au><au>Kitazume, Shinobu</au><au>Angata, Takashi</au><au>Fujinawa, Reiko</au><au>Ohtsubo, Kazuaki</au><au>Miyoshi, Eiji</au><au>Taniguchi, Naoyuki</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Simultaneous determination of nucleotide sugars with ion-pair reversed-phase HPLC</atitle><jtitle>Glycobiology (Oxford)</jtitle><addtitle>Glycobiology</addtitle><date>2010-07-01</date><risdate>2010</risdate><volume>20</volume><issue>7</issue><spage>865</spage><epage>871</epage><pages>865-871</pages><issn>0959-6658</issn><eissn>1460-2423</eissn><abstract>Nucleotide sugars are important in determining cell surface glycoprotein glycosylation, which can modulate cellular properties such as growth and arrest. We have developed a conventional HPLC method for simultaneous determination of nucleotide sugars. A mixture of nucleotide sugars (CMP-NeuAc, UDP-Gal, UDP-Glc, UDP-GalNAc, UDP-GlcNAc, GDP-Man, GDP-Fuc and UDP-GlcUA) and relevant nucleotides were perfectly separated in an optimized ion-pair reversed-phase mode using Inertsil ODS-4 and ODS-3 columns. The newly developed method enabled us to determine the nucleotide sugars in cellular extracts from 1 x 10⁶ cells in a single run. We applied this method to characterize nucleotide sugar levels in breast and pancreatic cancer cell lines and revealed that the abundance of UDP-GlcNAc, UDP-GalNAc, UDP-GlcUA and GDP-Fuc were a cell-type-specific feature. To determine the physiological significance of changes in nucleotide sugar levels, we analyzed their changes by glucose deprivation and found that the determination of nucleotide sugar levels provided us with valuable information with respect to studying the overview of cellular glycosylation status.</abstract><cop>England</cop><pub>Oxford University Press</pub><pmid>20371511</pmid><doi>10.1093/glycob/cwq044</doi><tpages>7</tpages><oa>free_for_read</oa></addata></record> |
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| source | MEDLINE; Oxford University Press Journals All Titles (1996-Current); EZB-FREE-00999 freely available EZB journals; Alma/SFX Local Collection |
| subjects | Carbohydrates - chemistry Cell Line, Tumor Chromatography, High Pressure Liquid - methods Chromatography, Ion Exchange - methods Glycosylation Humans ion-pair RP-HPLC Membrane Glycoproteins - chemistry Membrane Glycoproteins - metabolism nucleotide sugar Nucleotides - chemistry Nucleotides - metabolism Reproducibility of Results Sensitivity and Specificity |
| title | Simultaneous determination of nucleotide sugars with ion-pair reversed-phase HPLC |
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