Detection and Characterization of Low Abundance Glycopeptides Via Higher-Energy C-Trap Dissociation and Orbitrap Mass Analysis
Broad-scale mass spectrometric analyses of glycopeptides are constrained by the considerable complexity inherent to glycoproteomics, and techniques are still being actively developed to address the associated analytical difficulties. Here we apply Orbitrap mass analysis and higher-energy C-trap diss...
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| Veröffentlicht in: | Journal of the American Society for Mass Spectrometry 2012-01, Vol.23 (1), p.124-140 |
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| creator | Hart-Smith, Gene Raftery, Mark J. |
| description | Broad-scale mass spectrometric analyses of glycopeptides are constrained by the considerable complexity inherent to glycoproteomics, and techniques are still being actively developed to address the associated analytical difficulties. Here we apply Orbitrap mass analysis and higher-energy C-trap dissociation (HCD) to facilitate detailed insights into the compositions and heterogeneity of complex mixtures of low abundance glycopeptides. By generating diagnostic oxonium product ions at mass measurement errors of |
| doi_str_mv | 10.1007/s13361-011-0273-y |
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N-
acetylhexosamine (HexNAc) ions (Y1 ions) can be readily achieved at <5 ppm mass measurement errors. These data allow base peptide sequences and glycan compositional information to be attained with high confidence, even for glycopeptides that produce weak precursor ion signals and/or low quality MS/MS spectra. The glycopeptides characterized from low fmol abundances using these methods allow two previously unreported glycosylation sites on the
Gallus gallus
protein ovoglycoprotein (amino acids 82 and 90) to be confirmed; considerable glycan heterogeneities at amino acid 90 of ovoglycoprotein, and amino acids 34 and 77 of
Gallus gallus
ovomucoid are also revealed.</description><identifier>ISSN: 1044-0305</identifier><identifier>EISSN: 1879-1123</identifier><identifier>DOI: 10.1007/s13361-011-0273-y</identifier><identifier>PMID: 22083589</identifier><language>eng</language><publisher>New York: Springer-Verlag</publisher><subject>Abundance ; Amino Acid Sequence ; Amino acids ; Analytical Chemistry ; Analytical, structural and metabolic biochemistry ; Animals ; Antibiotics ; Bioinformatics ; Biological and medical sciences ; Biotechnology ; Cattle ; Chemistry ; Chemistry and Materials Science ; Chickens ; Complexity ; Diagnostic systems ; Energy of dissociation ; Fundamental and applied biological sciences. Psychology ; General aspects, investigation methods ; Glycan ; Glycopeptides ; Glycopeptides - analysis ; Glycopeptides - chemistry ; Glycoproteins ; Ions ; Ions - chemistry ; Limit of Detection ; Mass spectrometry ; Molecular Sequence Data ; Organic Chemistry ; Peptide Fragments - chemistry ; Peptides ; Polysaccharides - chemistry ; Proteins ; Proteins - chemistry ; Proteomics ; Research Article ; Sequences ; Spectrometry ; Tandem Mass Spectrometry - methods ; Trypsin - chemistry</subject><ispartof>Journal of the American Society for Mass Spectrometry, 2012-01, Vol.23 (1), p.124-140</ispartof><rights>American Society for Mass Spectrometry 2011</rights><rights>2015 INIST-CNRS</rights><rights>American Society for Mass Spectrometry, 2011</rights><rights>Journal of The American Society for Mass Spectrometry is a copyright of Springer, 2012.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c510t-2474fd81ba284ff9f7fbde9499de17d4550a86f165cf84b891f8b33951c070133</citedby><cites>FETCH-LOGICAL-c510t-2474fd81ba284ff9f7fbde9499de17d4550a86f165cf84b891f8b33951c070133</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://link.springer.com/content/pdf/10.1007/s13361-011-0273-y$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1007/s13361-011-0273-y$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>314,776,780,27901,27902,41464,42533,51294</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=25556390$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/22083589$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Hart-Smith, Gene</creatorcontrib><creatorcontrib>Raftery, Mark J.</creatorcontrib><title>Detection and Characterization of Low Abundance Glycopeptides Via Higher-Energy C-Trap Dissociation and Orbitrap Mass Analysis</title><title>Journal of the American Society for Mass Spectrometry</title><addtitle>J. Am. Soc. Mass Spectrom</addtitle><addtitle>J Am Soc Mass Spectrom</addtitle><description>Broad-scale mass spectrometric analyses of glycopeptides are constrained by the considerable complexity inherent to glycoproteomics, and techniques are still being actively developed to address the associated analytical difficulties. Here we apply Orbitrap mass analysis and higher-energy C-trap dissociation (HCD) to facilitate detailed insights into the compositions and heterogeneity of complex mixtures of low abundance glycopeptides. By generating diagnostic oxonium product ions at mass measurement errors of <5 ppm, highly selective glycopeptide precursor ion detections are made at sub-fmol limits of detection: analyses of proteolytic digests of a hen egg glycoprotein mixture detect 88 previously uncharacterized glycopeptides from 666 precursor ions selected for MS/MS, with only one false positive due to co-fragmentation of a non-glycosylated peptide with a glycopeptide. We also demonstrate that by (1) identifying multiple series of glycoforms using high mass accuracy single stage MS spectra, and (2) performing product ion scans at optimized HCD collision energies, the identification of peptide +
N-
acetylhexosamine (HexNAc) ions (Y1 ions) can be readily achieved at <5 ppm mass measurement errors. These data allow base peptide sequences and glycan compositional information to be attained with high confidence, even for glycopeptides that produce weak precursor ion signals and/or low quality MS/MS spectra. The glycopeptides characterized from low fmol abundances using these methods allow two previously unreported glycosylation sites on the
Gallus gallus
protein ovoglycoprotein (amino acids 82 and 90) to be confirmed; considerable glycan heterogeneities at amino acid 90 of ovoglycoprotein, and amino acids 34 and 77 of
Gallus gallus
ovomucoid are also revealed.</description><subject>Abundance</subject><subject>Amino Acid Sequence</subject><subject>Amino acids</subject><subject>Analytical Chemistry</subject><subject>Analytical, structural and metabolic biochemistry</subject><subject>Animals</subject><subject>Antibiotics</subject><subject>Bioinformatics</subject><subject>Biological and medical sciences</subject><subject>Biotechnology</subject><subject>Cattle</subject><subject>Chemistry</subject><subject>Chemistry and Materials Science</subject><subject>Chickens</subject><subject>Complexity</subject><subject>Diagnostic systems</subject><subject>Energy of dissociation</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>General aspects, investigation methods</subject><subject>Glycan</subject><subject>Glycopeptides</subject><subject>Glycopeptides - analysis</subject><subject>Glycopeptides - chemistry</subject><subject>Glycoproteins</subject><subject>Ions</subject><subject>Ions - chemistry</subject><subject>Limit of Detection</subject><subject>Mass spectrometry</subject><subject>Molecular Sequence Data</subject><subject>Organic Chemistry</subject><subject>Peptide Fragments - chemistry</subject><subject>Peptides</subject><subject>Polysaccharides - chemistry</subject><subject>Proteins</subject><subject>Proteins - chemistry</subject><subject>Proteomics</subject><subject>Research Article</subject><subject>Sequences</subject><subject>Spectrometry</subject><subject>Tandem Mass Spectrometry - methods</subject><subject>Trypsin - chemistry</subject><issn>1044-0305</issn><issn>1879-1123</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2012</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>8G5</sourceid><sourceid>BENPR</sourceid><sourceid>GUQSH</sourceid><sourceid>M2O</sourceid><recordid>eNp1kU1v1DAQhi0EoqXwA7ggSwhxCngcO7GPq-0X0qJeClfLceytq2wSPIlQOPDb62WXgpA4jDzyPPPOaF5CXgP7AIzVHxHKsoKCQQ5el8XyhJyCqnUBwMunOWdCFKxk8oS8QLxnDGqm6-fkhHOmSqn0Kfl57ifvpjj01PYtXd_ZZN3kU_xhf30OgW6G73TVzH1re-fpVbe4YfTjFFuP9Gu09Dpu73wqLnqftgtdF7fJjvQ8Ig4u2kflm9TEaV_5bBHpqrfdghFfkmfBduhfHd8z8uXy4nZ9XWxurj6tV5vCSWBTwUUtQqugsVyJEHSoQ9N6LbRuPdStkJJZVQWopAtKNEpDUE1ZagmO1Sxf6Yy8P-iOafg2e5zMLqLzXWd7P8xoNHDBNSieybf_kPfDnPK6aCDrMVlJqDIFB8qlATH5YMYUdzYtBpjZe2MO3pjsjdl7Y5bc8-aoPDc73z52_DYjA--OgEVnu5DywSP-4aSUValZ5viBw1zqtz79teJ_pz8Au36m-Q</recordid><startdate>20120101</startdate><enddate>20120101</enddate><creator>Hart-Smith, Gene</creator><creator>Raftery, Mark J.</creator><general>Springer-Verlag</general><general>Elsevier</general><general>Springer Nature B.V</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>3V.</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FE</scope><scope>8FG</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>8G5</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>ARAPS</scope><scope>AZQEC</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>GUQSH</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>M0S</scope><scope>M1P</scope><scope>M2O</scope><scope>MBDVC</scope><scope>P5Z</scope><scope>P62</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>Q9U</scope><scope>7X8</scope></search><sort><creationdate>20120101</creationdate><title>Detection and Characterization of Low Abundance Glycopeptides Via Higher-Energy C-Trap Dissociation and Orbitrap Mass Analysis</title><author>Hart-Smith, Gene ; Raftery, Mark J.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c510t-2474fd81ba284ff9f7fbde9499de17d4550a86f165cf84b891f8b33951c070133</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2012</creationdate><topic>Abundance</topic><topic>Amino Acid Sequence</topic><topic>Amino acids</topic><topic>Analytical Chemistry</topic><topic>Analytical, structural and metabolic biochemistry</topic><topic>Animals</topic><topic>Antibiotics</topic><topic>Bioinformatics</topic><topic>Biological and medical sciences</topic><topic>Biotechnology</topic><topic>Cattle</topic><topic>Chemistry</topic><topic>Chemistry and Materials Science</topic><topic>Chickens</topic><topic>Complexity</topic><topic>Diagnostic systems</topic><topic>Energy of dissociation</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>General aspects, investigation methods</topic><topic>Glycan</topic><topic>Glycopeptides</topic><topic>Glycopeptides - analysis</topic><topic>Glycopeptides - chemistry</topic><topic>Glycoproteins</topic><topic>Ions</topic><topic>Ions - chemistry</topic><topic>Limit of Detection</topic><topic>Mass spectrometry</topic><topic>Molecular Sequence Data</topic><topic>Organic Chemistry</topic><topic>Peptide Fragments - chemistry</topic><topic>Peptides</topic><topic>Polysaccharides - chemistry</topic><topic>Proteins</topic><topic>Proteins - chemistry</topic><topic>Proteomics</topic><topic>Research Article</topic><topic>Sequences</topic><topic>Spectrometry</topic><topic>Tandem Mass Spectrometry - methods</topic><topic>Trypsin - chemistry</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Hart-Smith, Gene</creatorcontrib><creatorcontrib>Raftery, Mark J.</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Technology Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>Research Library (Alumni Edition)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>Advanced Technologies & Aerospace Collection</collection><collection>ProQuest Central Essentials</collection><collection>ProQuest Central</collection><collection>Technology Collection</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>Research Library Prep</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Research Library</collection><collection>Research Library (Corporate)</collection><collection>Advanced Technologies & Aerospace Database</collection><collection>ProQuest Advanced Technologies & Aerospace Collection</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central China</collection><collection>ProQuest Central Basic</collection><collection>MEDLINE - Academic</collection><jtitle>Journal of the American Society for Mass Spectrometry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Hart-Smith, Gene</au><au>Raftery, Mark J.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Detection and Characterization of Low Abundance Glycopeptides Via Higher-Energy C-Trap Dissociation and Orbitrap Mass Analysis</atitle><jtitle>Journal of the American Society for Mass Spectrometry</jtitle><stitle>J. Am. Soc. Mass Spectrom</stitle><addtitle>J Am Soc Mass Spectrom</addtitle><date>2012-01-01</date><risdate>2012</risdate><volume>23</volume><issue>1</issue><spage>124</spage><epage>140</epage><pages>124-140</pages><issn>1044-0305</issn><eissn>1879-1123</eissn><abstract>Broad-scale mass spectrometric analyses of glycopeptides are constrained by the considerable complexity inherent to glycoproteomics, and techniques are still being actively developed to address the associated analytical difficulties. Here we apply Orbitrap mass analysis and higher-energy C-trap dissociation (HCD) to facilitate detailed insights into the compositions and heterogeneity of complex mixtures of low abundance glycopeptides. By generating diagnostic oxonium product ions at mass measurement errors of <5 ppm, highly selective glycopeptide precursor ion detections are made at sub-fmol limits of detection: analyses of proteolytic digests of a hen egg glycoprotein mixture detect 88 previously uncharacterized glycopeptides from 666 precursor ions selected for MS/MS, with only one false positive due to co-fragmentation of a non-glycosylated peptide with a glycopeptide. We also demonstrate that by (1) identifying multiple series of glycoforms using high mass accuracy single stage MS spectra, and (2) performing product ion scans at optimized HCD collision energies, the identification of peptide +
N-
acetylhexosamine (HexNAc) ions (Y1 ions) can be readily achieved at <5 ppm mass measurement errors. These data allow base peptide sequences and glycan compositional information to be attained with high confidence, even for glycopeptides that produce weak precursor ion signals and/or low quality MS/MS spectra. The glycopeptides characterized from low fmol abundances using these methods allow two previously unreported glycosylation sites on the
Gallus gallus
protein ovoglycoprotein (amino acids 82 and 90) to be confirmed; considerable glycan heterogeneities at amino acid 90 of ovoglycoprotein, and amino acids 34 and 77 of
Gallus gallus
ovomucoid are also revealed.</abstract><cop>New York</cop><pub>Springer-Verlag</pub><pmid>22083589</pmid><doi>10.1007/s13361-011-0273-y</doi><tpages>17</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Abundance Amino Acid Sequence Amino acids Analytical Chemistry Analytical, structural and metabolic biochemistry Animals Antibiotics Bioinformatics Biological and medical sciences Biotechnology Cattle Chemistry Chemistry and Materials Science Chickens Complexity Diagnostic systems Energy of dissociation Fundamental and applied biological sciences. Psychology General aspects, investigation methods Glycan Glycopeptides Glycopeptides - analysis Glycopeptides - chemistry Glycoproteins Ions Ions - chemistry Limit of Detection Mass spectrometry Molecular Sequence Data Organic Chemistry Peptide Fragments - chemistry Peptides Polysaccharides - chemistry Proteins Proteins - chemistry Proteomics Research Article Sequences Spectrometry Tandem Mass Spectrometry - methods Trypsin - chemistry |
| title | Detection and Characterization of Low Abundance Glycopeptides Via Higher-Energy C-Trap Dissociation and Orbitrap Mass Analysis |
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