Novel and cost-effective 6-plex isobaric tagging reagent, DiART, is effective for identification and relative quantification of complex protein mixtures using PQD fragmentation
Deuterium isobaric Amine Reactive Tag (DiART) reagents facilitate relative quantification during proteomic analysis in a functionally similar manner to commercially available isobaric tag for relative and absolute quantitation (iTRAQ) and tandem mass tag (TMT) reagents. In contrast to iTRAQ and TMT,...
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| Veröffentlicht in: | Journal of mass spectrometry. 2013-09, Vol.48 (9), p.1032-1041 |
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| creator | Ramsubramaniam, Nikhil Tao, Feng Li, Shuwei Marten, Mark R. |
| description | Deuterium isobaric Amine Reactive Tag (DiART) reagents facilitate relative quantification during proteomic analysis in a functionally similar manner to commercially available isobaric tag for relative and absolute quantitation (iTRAQ) and tandem mass tag (TMT) reagents. In contrast to iTRAQ and TMT, DiART reagents incorporate deuterium isotopes which significantly reduce the number of required synthesis steps and hence have potential to significantly reduce reagent production cost. We examined the capability of DiART for performing quantitative proteomic experiments using a linear ion‐trap mass spectrometer with Pulsed Q Dissociation (PQD) fragmentation. Using a synthetic peptide tagged with DiART reagent, we observed a precise mass shift of 144.79 Da on the triply charged precursor ion, which shows complete derivatization of the N‐terminus and ε‐amino group of lysine. A DiART tagged tryptic digest of bovine serum albumin showed a sequence coverage of 57.99% which was very comparable to that showed by iTRAQ, 54.77%. Furthermore, a ten protein mixture tagged with DiART reagents and mixed in 1:1:1:1:1:1 exhibited |
| doi_str_mv | 10.1002/jms.3249 |
| format | Article |
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In contrast to iTRAQ and TMT, DiART reagents incorporate deuterium isotopes which significantly reduce the number of required synthesis steps and hence have potential to significantly reduce reagent production cost. We examined the capability of DiART for performing quantitative proteomic experiments using a linear ion‐trap mass spectrometer with Pulsed Q Dissociation (PQD) fragmentation. Using a synthetic peptide tagged with DiART reagent, we observed a precise mass shift of 144.79 Da on the triply charged precursor ion, which shows complete derivatization of the N‐terminus and ε‐amino group of lysine. A DiART tagged tryptic digest of bovine serum albumin showed a sequence coverage of 57.99% which was very comparable to that showed by iTRAQ, 54.77%. Furthermore, a ten protein mixture tagged with DiART reagents and mixed in 1:1:1:1:1:1 exhibited < 15% error, whereas a linear trend (slope of 1.085) was observed when tagged proteins were mixed in the ratio 2:1:2:4:10:14 and plotted against theoretical ratios. Finally, when complex cell‐wall protein mixtures from the model fungus A. nidulans were tagged with DiART reagents and mixed in different ratios, they exhibited similar trends. We conclude that DiART reagents are capable of performing quantitative proteomic experiments using PQD on a linear ion trap mass spectrometer. Copyright © 2013 John Wiley & Sons, Ltd.</description><identifier>ISSN: 1076-5174</identifier><identifier>EISSN: 1096-9888</identifier><identifier>DOI: 10.1002/jms.3249</identifier><identifier>PMID: 24078244</identifier><language>eng</language><publisher>England: Blackwell Publishing Ltd</publisher><subject>Amines - chemistry ; Amino Acid Sequence ; Animals ; Aspergillus nidulans ; Aspergillus nidulans - chemistry ; Cattle ; Cell Wall - chemistry ; cell-wall proteins ; Chickens ; DiART ; Fungal Proteins - analysis ; Horses ; Indicators and Reagents ; linear ion trap ; Molecular Sequence Data ; PQD ; Proteins - analysis ; Proteomics - economics ; Proteomics - methods ; Spectrometry, Mass, Electrospray Ionization - methods ; Tandem Mass Spectrometry - methods</subject><ispartof>Journal of mass spectrometry., 2013-09, Vol.48 (9), p.1032-1041</ispartof><rights>Copyright © 2013 John Wiley & Sons, Ltd.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c4209-1d826acbf6763a59e20ce03e428c728624b4b2514de2dcb8c51f9a57a43a62273</citedby><cites>FETCH-LOGICAL-c4209-1d826acbf6763a59e20ce03e428c728624b4b2514de2dcb8c51f9a57a43a62273</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1002%2Fjms.3249$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1002%2Fjms.3249$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>314,780,784,1417,27924,27925,45574,45575</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/24078244$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Ramsubramaniam, Nikhil</creatorcontrib><creatorcontrib>Tao, Feng</creatorcontrib><creatorcontrib>Li, Shuwei</creatorcontrib><creatorcontrib>Marten, Mark R.</creatorcontrib><title>Novel and cost-effective 6-plex isobaric tagging reagent, DiART, is effective for identification and relative quantification of complex protein mixtures using PQD fragmentation</title><title>Journal of mass spectrometry.</title><addtitle>J. Mass Spectrom</addtitle><description>Deuterium isobaric Amine Reactive Tag (DiART) reagents facilitate relative quantification during proteomic analysis in a functionally similar manner to commercially available isobaric tag for relative and absolute quantitation (iTRAQ) and tandem mass tag (TMT) reagents. In contrast to iTRAQ and TMT, DiART reagents incorporate deuterium isotopes which significantly reduce the number of required synthesis steps and hence have potential to significantly reduce reagent production cost. We examined the capability of DiART for performing quantitative proteomic experiments using a linear ion‐trap mass spectrometer with Pulsed Q Dissociation (PQD) fragmentation. Using a synthetic peptide tagged with DiART reagent, we observed a precise mass shift of 144.79 Da on the triply charged precursor ion, which shows complete derivatization of the N‐terminus and ε‐amino group of lysine. A DiART tagged tryptic digest of bovine serum albumin showed a sequence coverage of 57.99% which was very comparable to that showed by iTRAQ, 54.77%. Furthermore, a ten protein mixture tagged with DiART reagents and mixed in 1:1:1:1:1:1 exhibited < 15% error, whereas a linear trend (slope of 1.085) was observed when tagged proteins were mixed in the ratio 2:1:2:4:10:14 and plotted against theoretical ratios. Finally, when complex cell‐wall protein mixtures from the model fungus A. nidulans were tagged with DiART reagents and mixed in different ratios, they exhibited similar trends. We conclude that DiART reagents are capable of performing quantitative proteomic experiments using PQD on a linear ion trap mass spectrometer. Copyright © 2013 John Wiley & Sons, Ltd.</description><subject>Amines - chemistry</subject><subject>Amino Acid Sequence</subject><subject>Animals</subject><subject>Aspergillus nidulans</subject><subject>Aspergillus nidulans - chemistry</subject><subject>Cattle</subject><subject>Cell Wall - chemistry</subject><subject>cell-wall proteins</subject><subject>Chickens</subject><subject>DiART</subject><subject>Fungal Proteins - analysis</subject><subject>Horses</subject><subject>Indicators and Reagents</subject><subject>linear ion trap</subject><subject>Molecular Sequence Data</subject><subject>PQD</subject><subject>Proteins - analysis</subject><subject>Proteomics - economics</subject><subject>Proteomics - methods</subject><subject>Spectrometry, Mass, Electrospray Ionization - methods</subject><subject>Tandem Mass Spectrometry - methods</subject><issn>1076-5174</issn><issn>1096-9888</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2013</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqNkUtv1DAUhSMEomVA4hcgS2xYNMV2_IiXVQudonYotDx2luPcRB7ymNpJO_1X_ESc6VAqJCRW19L57jnXOknykuB9gjF9u2zDfkaZepTsEqxEqvI8fzy9pUg5kWwneRbCEmOsFBNPkx3KsMwpY7vJz0V_DQ0yXYlsH4YUqgrs4K4BiXTVwBq50BfGO4sGU9euq5EHU0M37KEjd_D5ci8C6M9S1Xvkyii7ylkzuL7bWHtozEa_Gs1Dra9iarvJWfl-ANeh1q2H0UNAY5jSzj8docqbuo2em53nyZPKNAFebOcs-fL-3eXhPD39eHxyeHCaWkaxSkmZU2FsUQkpMsMVUGwBZ8BobiXNBWUFKygnrARa2iK3nFTKcGlYZgSlMpslb-5842FXI4RBty5YaBrTQT8GTRjLJRNC8v9As5zLDMc5S17_hS770XfxIxMlCVdcPci2vg_BQ6VX3rXG32qC9VS4joXrqfCIvtoajkUL5T34u-EIpHfAjWvg9p9G-sPZxdZwy7swwPqeN_6HFjKTXH9bHOtz9v3i69l8rhfZL9bCxag</recordid><startdate>201309</startdate><enddate>201309</enddate><creator>Ramsubramaniam, Nikhil</creator><creator>Tao, Feng</creator><creator>Li, Shuwei</creator><creator>Marten, Mark R.</creator><general>Blackwell Publishing Ltd</general><general>Wiley Subscription Services, Inc</general><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>7QF</scope><scope>7QO</scope><scope>7QP</scope><scope>7QQ</scope><scope>7QR</scope><scope>7SC</scope><scope>7SE</scope><scope>7SP</scope><scope>7SR</scope><scope>7TA</scope><scope>7TB</scope><scope>7TK</scope><scope>7U5</scope><scope>7U7</scope><scope>8BQ</scope><scope>8FD</scope><scope>C1K</scope><scope>F1W</scope><scope>F28</scope><scope>FR3</scope><scope>H8D</scope><scope>H8G</scope><scope>H97</scope><scope>JG9</scope><scope>JQ2</scope><scope>K9.</scope><scope>KR7</scope><scope>L.G</scope><scope>L7M</scope><scope>L~C</scope><scope>L~D</scope><scope>P64</scope><scope>7X8</scope><scope>M7N</scope></search><sort><creationdate>201309</creationdate><title>Novel and cost-effective 6-plex isobaric tagging reagent, DiART, is effective for identification and relative quantification of complex protein mixtures using PQD fragmentation</title><author>Ramsubramaniam, Nikhil ; 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Mass Spectrom</addtitle><date>2013-09</date><risdate>2013</risdate><volume>48</volume><issue>9</issue><spage>1032</spage><epage>1041</epage><pages>1032-1041</pages><issn>1076-5174</issn><eissn>1096-9888</eissn><abstract>Deuterium isobaric Amine Reactive Tag (DiART) reagents facilitate relative quantification during proteomic analysis in a functionally similar manner to commercially available isobaric tag for relative and absolute quantitation (iTRAQ) and tandem mass tag (TMT) reagents. In contrast to iTRAQ and TMT, DiART reagents incorporate deuterium isotopes which significantly reduce the number of required synthesis steps and hence have potential to significantly reduce reagent production cost. We examined the capability of DiART for performing quantitative proteomic experiments using a linear ion‐trap mass spectrometer with Pulsed Q Dissociation (PQD) fragmentation. Using a synthetic peptide tagged with DiART reagent, we observed a precise mass shift of 144.79 Da on the triply charged precursor ion, which shows complete derivatization of the N‐terminus and ε‐amino group of lysine. A DiART tagged tryptic digest of bovine serum albumin showed a sequence coverage of 57.99% which was very comparable to that showed by iTRAQ, 54.77%. Furthermore, a ten protein mixture tagged with DiART reagents and mixed in 1:1:1:1:1:1 exhibited < 15% error, whereas a linear trend (slope of 1.085) was observed when tagged proteins were mixed in the ratio 2:1:2:4:10:14 and plotted against theoretical ratios. Finally, when complex cell‐wall protein mixtures from the model fungus A. nidulans were tagged with DiART reagents and mixed in different ratios, they exhibited similar trends. We conclude that DiART reagents are capable of performing quantitative proteomic experiments using PQD on a linear ion trap mass spectrometer. Copyright © 2013 John Wiley & Sons, Ltd.</abstract><cop>England</cop><pub>Blackwell Publishing Ltd</pub><pmid>24078244</pmid><doi>10.1002/jms.3249</doi><tpages>10</tpages></addata></record> |
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| ispartof | Journal of mass spectrometry., 2013-09, Vol.48 (9), p.1032-1041 |
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| subjects | Amines - chemistry Amino Acid Sequence Animals Aspergillus nidulans Aspergillus nidulans - chemistry Cattle Cell Wall - chemistry cell-wall proteins Chickens DiART Fungal Proteins - analysis Horses Indicators and Reagents linear ion trap Molecular Sequence Data PQD Proteins - analysis Proteomics - economics Proteomics - methods Spectrometry, Mass, Electrospray Ionization - methods Tandem Mass Spectrometry - methods |
| title | Novel and cost-effective 6-plex isobaric tagging reagent, DiART, is effective for identification and relative quantification of complex protein mixtures using PQD fragmentation |
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