Rapid Mass Spectrometric Peptide Sequencing and Mass Matching for Characterization of Human Melanoma Proteins Isolated by Two-Dimensional PAGE
We report a general mass spectrometric approach for the rapid identification and characterization of proteins isolated by preparative two-dimensional polyacrylamide gel electrophoresis. This method possesses the inherent power to detect and structurally characterize covalent modifications. Absolute...
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| Veröffentlicht in: | Proceedings of the National Academy of Sciences - PNAS 1995-05, Vol.92 (11), p.5072-5076 |
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| container_title | Proceedings of the National Academy of Sciences - PNAS |
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| creator | Clauser, Karl R. Hall, Steven C. Smith, Diana M. Webb, James W. Andrews, Lori E. Tran, Huu M. Epstein, Lois B. Burlingame, Alma L. |
| description | We report a general mass spectrometric approach for the rapid identification and characterization of proteins isolated by preparative two-dimensional polyacrylamide gel electrophoresis. This method possesses the inherent power to detect and structurally characterize covalent modifications. Absolute sensitivities of matrix-assisted laser desorption ionization and high-energy collision-induced dissociation tandem mass spectrometry are exploited to determine the mass and sequence of subpicomole sample quantities of tryptic peptides. These data permit mass matching and sequence homology searching of computerized peptide mass and protein sequence data bases for known proteins and design of oligonucleotide probes for cloning unknown proteins. We have identified 11 proteins in lysates of human A375 melanoma cells, including: α-enolase, cytokeratin, stathmin, protein disulfide isomerase, tropomyosin, Cu/Zn superoxide dismutase, nucleoside diphosphate kinase A, galaptin, and triosephosphate isomerase. We have characterized several post-translational modifications and chemical modifications that may result from electrophoresis or subsequent sample processing steps. Detection of comigrating and covalently modified proteins illustrates the necessity of peptide sequencing and the advantages of tandem mass spectrometry to reliably and unambiguously establish the identity of each protein. This technology paves the way for studies of cell-type dependent gene expression and studies of large suites of cellular proteins with unprecedented speed and rigor to provide information complementary to the ongoing Human Genome Project. |
| doi_str_mv | 10.1073/pnas.92.11.5072 |
| format | Article |
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This method possesses the inherent power to detect and structurally characterize covalent modifications. Absolute sensitivities of matrix-assisted laser desorption ionization and high-energy collision-induced dissociation tandem mass spectrometry are exploited to determine the mass and sequence of subpicomole sample quantities of tryptic peptides. These data permit mass matching and sequence homology searching of computerized peptide mass and protein sequence data bases for known proteins and design of oligonucleotide probes for cloning unknown proteins. We have identified 11 proteins in lysates of human A375 melanoma cells, including: α-enolase, cytokeratin, stathmin, protein disulfide isomerase, tropomyosin, Cu/Zn superoxide dismutase, nucleoside diphosphate kinase A, galaptin, and triosephosphate isomerase. We have characterized several post-translational modifications and chemical modifications that may result from electrophoresis or subsequent sample processing steps. Detection of comigrating and covalently modified proteins illustrates the necessity of peptide sequencing and the advantages of tandem mass spectrometry to reliably and unambiguously establish the identity of each protein. This technology paves the way for studies of cell-type dependent gene expression and studies of large suites of cellular proteins with unprecedented speed and rigor to provide information complementary to the ongoing Human Genome Project.</description><identifier>ISSN: 0027-8424</identifier><identifier>EISSN: 1091-6490</identifier><identifier>DOI: 10.1073/pnas.92.11.5072</identifier><identifier>PMID: 7761450</identifier><language>eng</language><publisher>United States: National Academy of Sciences of the United States of America</publisher><subject>Amino Acid Sequence ; Average linear density ; Biochemistry ; Cell Line ; Chromatography, High Pressure Liquid ; Databases, Factual ; Electrophoresis ; Enzymes - chemistry ; Enzymes - isolation & purification ; Gels ; Humans ; Isoelectric Focusing ; Keratins ; Mass spectrometers ; Mass Spectrometry - methods ; Mass spectroscopy ; Melanoma ; Melanoma - chemistry ; Molecular Sequence Data ; Molecular Weight ; Neoplasm Proteins - chemistry ; Neoplasm Proteins - isolation & purification ; Peptides ; Peptides - chemistry ; Proteins ; Proteins - chemistry ; Proteins - isolation & purification ; Scientific imaging ; Sequence Homology, Amino Acid ; Sequencing ; Tumor Cells, Cultured ; Tumors</subject><ispartof>Proceedings of the National Academy of Sciences - PNAS, 1995-05, Vol.92 (11), p.5072-5076</ispartof><rights>Copyright 1995 The National Academy of Sciences of the United States of America</rights><rights>Copyright National Academy of Sciences May 23, 1995</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c5302-45ceb9c1c8758b12b4860ca92568b44c6a6bed59d272ede7943d0ded069d33c93</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Uhttp://www.pnas.org/content/92/11.cover.gif</thumbnail><linktopdf>$$Uhttps://www.jstor.org/stable/pdf/2367673$$EPDF$$P50$$Gjstor$$H</linktopdf><linktohtml>$$Uhttps://www.jstor.org/stable/2367673$$EHTML$$P50$$Gjstor$$H</linktohtml><link.rule.ids>230,314,723,776,780,799,881,27901,27902,53766,53768,57992,58225</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/7761450$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Clauser, Karl R.</creatorcontrib><creatorcontrib>Hall, Steven C.</creatorcontrib><creatorcontrib>Smith, Diana M.</creatorcontrib><creatorcontrib>Webb, James W.</creatorcontrib><creatorcontrib>Andrews, Lori E.</creatorcontrib><creatorcontrib>Tran, Huu M.</creatorcontrib><creatorcontrib>Epstein, Lois B.</creatorcontrib><creatorcontrib>Burlingame, Alma L.</creatorcontrib><title>Rapid Mass Spectrometric Peptide Sequencing and Mass Matching for Characterization of Human Melanoma Proteins Isolated by Two-Dimensional PAGE</title><title>Proceedings of the National Academy of Sciences - PNAS</title><addtitle>Proc Natl Acad Sci U S A</addtitle><description>We report a general mass spectrometric approach for the rapid identification and characterization of proteins isolated by preparative two-dimensional polyacrylamide gel electrophoresis. This method possesses the inherent power to detect and structurally characterize covalent modifications. Absolute sensitivities of matrix-assisted laser desorption ionization and high-energy collision-induced dissociation tandem mass spectrometry are exploited to determine the mass and sequence of subpicomole sample quantities of tryptic peptides. These data permit mass matching and sequence homology searching of computerized peptide mass and protein sequence data bases for known proteins and design of oligonucleotide probes for cloning unknown proteins. We have identified 11 proteins in lysates of human A375 melanoma cells, including: α-enolase, cytokeratin, stathmin, protein disulfide isomerase, tropomyosin, Cu/Zn superoxide dismutase, nucleoside diphosphate kinase A, galaptin, and triosephosphate isomerase. We have characterized several post-translational modifications and chemical modifications that may result from electrophoresis or subsequent sample processing steps. Detection of comigrating and covalently modified proteins illustrates the necessity of peptide sequencing and the advantages of tandem mass spectrometry to reliably and unambiguously establish the identity of each protein. This technology paves the way for studies of cell-type dependent gene expression and studies of large suites of cellular proteins with unprecedented speed and rigor to provide information complementary to the ongoing Human Genome Project.</description><subject>Amino Acid Sequence</subject><subject>Average linear density</subject><subject>Biochemistry</subject><subject>Cell Line</subject><subject>Chromatography, High Pressure Liquid</subject><subject>Databases, Factual</subject><subject>Electrophoresis</subject><subject>Enzymes - chemistry</subject><subject>Enzymes - isolation & purification</subject><subject>Gels</subject><subject>Humans</subject><subject>Isoelectric Focusing</subject><subject>Keratins</subject><subject>Mass spectrometers</subject><subject>Mass Spectrometry - methods</subject><subject>Mass spectroscopy</subject><subject>Melanoma</subject><subject>Melanoma - chemistry</subject><subject>Molecular Sequence Data</subject><subject>Molecular Weight</subject><subject>Neoplasm Proteins - chemistry</subject><subject>Neoplasm Proteins - isolation & purification</subject><subject>Peptides</subject><subject>Peptides - chemistry</subject><subject>Proteins</subject><subject>Proteins - chemistry</subject><subject>Proteins - isolation & purification</subject><subject>Scientific imaging</subject><subject>Sequence Homology, Amino Acid</subject><subject>Sequencing</subject><subject>Tumor Cells, Cultured</subject><subject>Tumors</subject><issn>0027-8424</issn><issn>1091-6490</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1995</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkk1vEzEQhlcIVELhzAWQxQFOm9reD9sSlyqUtlIjIlrOlteeNI5214vtBcqP4DfjVUL4OMDJ0rzPO_I7M1n2lOA5waw4GXoV5oLOCZlXmNF72YxgQfK6FPh-NsOYspyXtHyYPQphizEWFcdH2RFjNSkrPMu-f1CDNWipQkDXA-joXQfRW41WMERrAF3DpxF6bftbpPo9uVRRb6bK2nm02CivdARvv6loXY_cGl2MnerRElrVu06hlXcRbB_QZXCtimBQc4duvrj8re2gD8mkWrQ6PT97nD1YqzbAk_17nH18d3azuMiv3p9fLk6vcl0VmOZlpaERmmjOKt4Q2pS8xloJWtW8KUtdq7oBUwlDGQUDTJSFwQYMroUpCi2K4-zNru8wNh0YDX30qpWDt53yd9IpK_9UeruRt-6zLAmvcLK_2tu9S9MJUXY2aGhTXHBjkIxRUXDG_wuSmuMqrSSBL_8Ct270aSxBUkxSZk6KBJ3sIO1dCB7Whw8TLKdzkNM5SEElIXI6h-R4_nvOA7_ff9Jf7_XJ-FP91UCux7aN8DUm8sU_yQQ82wHbEJ0_ELSoWc2K4gfnidR8</recordid><startdate>19950523</startdate><enddate>19950523</enddate><creator>Clauser, Karl R.</creator><creator>Hall, Steven C.</creator><creator>Smith, Diana M.</creator><creator>Webb, James W.</creator><creator>Andrews, Lori E.</creator><creator>Tran, Huu M.</creator><creator>Epstein, Lois B.</creator><creator>Burlingame, Alma L.</creator><general>National Academy of Sciences of the United States of America</general><general>National Acad Sciences</general><general>National Academy of Sciences</general><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>7QG</scope><scope>7QL</scope><scope>7QP</scope><scope>7QR</scope><scope>7SN</scope><scope>7SS</scope><scope>7T5</scope><scope>7TK</scope><scope>7TM</scope><scope>7TO</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>19950523</creationdate><title>Rapid Mass Spectrometric Peptide Sequencing and Mass Matching for Characterization of Human Melanoma Proteins Isolated by Two-Dimensional PAGE</title><author>Clauser, Karl R. ; 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This method possesses the inherent power to detect and structurally characterize covalent modifications. Absolute sensitivities of matrix-assisted laser desorption ionization and high-energy collision-induced dissociation tandem mass spectrometry are exploited to determine the mass and sequence of subpicomole sample quantities of tryptic peptides. These data permit mass matching and sequence homology searching of computerized peptide mass and protein sequence data bases for known proteins and design of oligonucleotide probes for cloning unknown proteins. We have identified 11 proteins in lysates of human A375 melanoma cells, including: α-enolase, cytokeratin, stathmin, protein disulfide isomerase, tropomyosin, Cu/Zn superoxide dismutase, nucleoside diphosphate kinase A, galaptin, and triosephosphate isomerase. We have characterized several post-translational modifications and chemical modifications that may result from electrophoresis or subsequent sample processing steps. Detection of comigrating and covalently modified proteins illustrates the necessity of peptide sequencing and the advantages of tandem mass spectrometry to reliably and unambiguously establish the identity of each protein. This technology paves the way for studies of cell-type dependent gene expression and studies of large suites of cellular proteins with unprecedented speed and rigor to provide information complementary to the ongoing Human Genome Project.</abstract><cop>United States</cop><pub>National Academy of Sciences of the United States of America</pub><pmid>7761450</pmid><doi>10.1073/pnas.92.11.5072</doi><tpages>5</tpages><oa>free_for_read</oa></addata></record> |
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| identifier | ISSN: 0027-8424 |
| ispartof | Proceedings of the National Academy of Sciences - PNAS, 1995-05, Vol.92 (11), p.5072-5076 |
| issn | 0027-8424 1091-6490 |
| language | eng |
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| source | Jstor Complete Legacy; MEDLINE; PubMed Central; Alma/SFX Local Collection; Free Full-Text Journals in Chemistry |
| subjects | Amino Acid Sequence Average linear density Biochemistry Cell Line Chromatography, High Pressure Liquid Databases, Factual Electrophoresis Enzymes - chemistry Enzymes - isolation & purification Gels Humans Isoelectric Focusing Keratins Mass spectrometers Mass Spectrometry - methods Mass spectroscopy Melanoma Melanoma - chemistry Molecular Sequence Data Molecular Weight Neoplasm Proteins - chemistry Neoplasm Proteins - isolation & purification Peptides Peptides - chemistry Proteins Proteins - chemistry Proteins - isolation & purification Scientific imaging Sequence Homology, Amino Acid Sequencing Tumor Cells, Cultured Tumors |
| title | Rapid Mass Spectrometric Peptide Sequencing and Mass Matching for Characterization of Human Melanoma Proteins Isolated by Two-Dimensional PAGE |
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