TMTpro reagents: a set of isobaric labeling mass tags enables simultaneous proteome-wide measurements across 16 samples
Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on an isobutyl-proline immonium ion reporter structure (TMTpro) is presented. These reagents have similar characteristics to existing tandem mas...
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| Published in | Nature methods Vol. 17; no. 4; pp. 399 - 404 |
|---|---|
| Main Authors | , , , , , , , , , , , , , , , |
| Format | Journal Article |
| Language | English |
| Published |
New York
Nature Publishing Group US
01.04.2020
Nature Publishing Group |
| Subjects | |
| Online Access | Get full text |
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| Abstract | Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on an isobutyl-proline immonium ion reporter structure (TMTpro) is presented. These reagents have similar characteristics to existing tandem mass tag reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides per protein) per replicate with an analysis time of only 1.1 h per proteome. Finally, we modified the thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20 µM staurosporine with five replicates. This assay identified and dose-stratified staurosporine binding to 228 cellular kinases in just one, 18-h experiment. TMTpro reagents allow complex experimental designs—all with essentially no missing values across the 16 samples and no loss in quantitative integrity.
A set of isobaric labeling reagents called TMTpro enables deep quantitative comparisons of proteome measurements across 16 samples. |
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| AbstractList | Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on an isobutyl-proline immonium ion reporter structure (TMTpro) is presented. These reagents have similar characteristics to existing tandem mass tag reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides per protein) per replicate with an analysis time of only 1.1 h per proteome. Finally, we modified the thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20 μM staurosporine with five replicates. This assay identified and dose-stratified staurosporine binding to 228 cellular kinases in just one, 18-h experiment. TMTpro reagents allow complex experimental designs—all with essentially no missing values across the 16 samples and no loss in quantitative integrity. Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on an isobutyl-proline immonium ion reporter structure (TMTpro) is presented. These reagents have similar characteristics to existing tandem mass tag reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides per protein) per replicate with an analysis time of only 1.1 h per proteome. Finally, we modified the thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20 µM staurosporine with five replicates. This assay identified and dose-stratified staurosporine binding to 228 cellular kinases in just one, 18-h experiment. TMTpro reagents allow complex experimental designs-all with essentially no missing values across the 16 samples and no loss in quantitative integrity. Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on an isobutyl-proline immonium ion reporter structure (TMTpro) is presented. These reagents have similar characteristics to existing tandem mass tag reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides per protein) per replicate with an analysis time of only 1.1 h per proteome. Finally, we modified the thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20 µM staurosporine with five replicates. This assay identified and dose-stratified staurosporine binding to 228 cellular kinases in just one, 18-h experiment. TMTpro reagents allow complex experimental designs—all with essentially no missing values across the 16 samples and no loss in quantitative integrity. A set of isobaric labeling reagents called TMTpro enables deep quantitative comparisons of proteome measurements across 16 samples. Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on an isobutyl-proline immonium ion reporter structure (TMTpro) is presented. These reagents have similar characteristics to existing tandem mass tag reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides per protein) per replicate with an analysis time of only 1.1 h per proteome. Finally, we modified the thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20 [micro]M staurosporine with five replicates. This assay identified and dose-stratified staurosporine binding to 228 cellular kinases in just one, 18-h experiment. TMTpro reagents allow complex experimental designs--all with essentially no missing values across the 16 samples and no loss in quantitative integrity. Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on an isobutyl-proline immonium ion reporter structure (TMTpro) is presented. These reagents have similar characteristics to existing tandem mass tag reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides per protein) per replicate with an analysis time of only 1.1 h per proteome. Finally, we modified the thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20 [micro]M staurosporine with five replicates. This assay identified and dose-stratified staurosporine binding to 228 cellular kinases in just one, 18-h experiment. TMTpro reagents allow complex experimental designs--all with essentially no missing values across the 16 samples and no loss in quantitative integrity. A set of isobaric labeling reagents called TMTpro enables deep quantitative comparisons of proteome measurements across 16 samples. Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on an isobutyl-proline immonium ion reporter structure (TMTpro) is presented. These reagents have similar characteristics to existing tandem mass tag reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides per protein) per replicate with an analysis time of only 1.1 h per proteome. Finally, we modified the thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20 µM staurosporine with five replicates. This assay identified and dose-stratified staurosporine binding to 228 cellular kinases in just one, 18-h experiment. TMTpro reagents allow complex experimental designs—all with essentially no missing values across the 16 samples and no loss in quantitative integrity.A set of isobaric labeling reagents called TMTpro enables deep quantitative comparisons of proteome measurements across 16 samples. |
| Audience | Academic |
| Author | Pike, Ian Kuhn, Karsten Bomgarden, Ryan D. Schweppe, Devin K. Etienne, Chris Viner, Rosa Gygi, Steven P. Rogers, John C. Van Vranken, Jonathan G. Li, Jiaming Huttlin, Edward L. Pontano Vaites, Laura Paulo, Joao A. Nandhikonda, Premchendar Robitaille, Aaron M. Thompson, Andrew H. |
| AuthorAffiliation | 3 Thermo Fisher Scientific, San Jose, CA, USA 2 Thermo Fisher Scientific, Rockford, IL, USA 4 Proteome Sciences, London, UK 1 Department of Cell Biology, Harvard Medical School, Boston, MA, USA |
| AuthorAffiliation_xml | – name: 1 Department of Cell Biology, Harvard Medical School, Boston, MA, USA – name: 2 Thermo Fisher Scientific, Rockford, IL, USA – name: 3 Thermo Fisher Scientific, San Jose, CA, USA – name: 4 Proteome Sciences, London, UK |
| Author_xml | – sequence: 1 givenname: Jiaming orcidid: 0000-0002-9065-6913 surname: Li fullname: Li, Jiaming organization: Department of Cell Biology, Harvard Medical School – sequence: 2 givenname: Jonathan G. surname: Van Vranken fullname: Van Vranken, Jonathan G. organization: Department of Cell Biology, Harvard Medical School – sequence: 3 givenname: Laura surname: Pontano Vaites fullname: Pontano Vaites, Laura organization: Department of Cell Biology, Harvard Medical School – sequence: 4 givenname: Devin K. surname: Schweppe fullname: Schweppe, Devin K. organization: Department of Cell Biology, Harvard Medical School – sequence: 5 givenname: Edward L. surname: Huttlin fullname: Huttlin, Edward L. organization: Department of Cell Biology, Harvard Medical School – sequence: 6 givenname: Chris surname: Etienne fullname: Etienne, Chris organization: Thermo Fisher Scientific – sequence: 7 givenname: Premchendar surname: Nandhikonda fullname: Nandhikonda, Premchendar organization: Thermo Fisher Scientific – sequence: 8 givenname: Rosa orcidid: 0000-0003-0550-5545 surname: Viner fullname: Viner, Rosa organization: Thermo Fisher Scientific – sequence: 9 givenname: Aaron M. surname: Robitaille fullname: Robitaille, Aaron M. organization: Thermo Fisher Scientific – sequence: 10 givenname: Andrew H. surname: Thompson fullname: Thompson, Andrew H. organization: Proteome Sciences – sequence: 11 givenname: Karsten surname: Kuhn fullname: Kuhn, Karsten organization: Proteome Sciences – sequence: 12 givenname: Ian surname: Pike fullname: Pike, Ian organization: Proteome Sciences – sequence: 13 givenname: Ryan D. surname: Bomgarden fullname: Bomgarden, Ryan D. organization: Thermo Fisher Scientific – sequence: 14 givenname: John C. surname: Rogers fullname: Rogers, John C. organization: Thermo Fisher Scientific – sequence: 15 givenname: Steven P. orcidid: 0000-0001-7626-0034 surname: Gygi fullname: Gygi, Steven P. email: Steven_Gygi@hms.harvard.edu organization: Department of Cell Biology, Harvard Medical School – sequence: 16 givenname: Joao A. orcidid: 0000-0002-4291-413X surname: Paulo fullname: Paulo, Joao A. email: Joao_Paulo@hms.harvard.edu organization: Department of Cell Biology, Harvard Medical School |
| BackLink | https://www.ncbi.nlm.nih.gov/pubmed/32203386$$D View this record in MEDLINE/PubMed |
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| ContentType | Journal Article |
| Copyright | The Author(s), under exclusive licence to Springer Nature America, Inc. 2020 COPYRIGHT 2020 Nature Publishing Group The Author(s), under exclusive licence to Springer Nature America, Inc. 2020. |
| Copyright_xml | – notice: The Author(s), under exclusive licence to Springer Nature America, Inc. 2020 – notice: COPYRIGHT 2020 Nature Publishing Group – notice: The Author(s), under exclusive licence to Springer Nature America, Inc. 2020. |
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| DOI | 10.1038/s41592-020-0781-4 |
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| Notes | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 J.L. prepared the cell line samples treated with Torin1 for MS analysis, performed the data analyses, prepared the figures and wrote the manuscript. J.G.V.V. prepared and conducted the PISA experiments and prepared associated figures. L.P.V. proposed the eight cell lines, then grew and treated the lines for Torin1 experiments and performed western blotting experiments. D.K.S. developed and implemented the real-time online searching tool. E.L.H. advised on data analyses. R.V. and A.M.R. provided additional reagent characterization and advice. C.E., P.N., R.D.B. and J.C.R. further characterized the TMTpro reagents, initiated the collaboration and provided input and oversight for the project. Reagents were conceived, developed, synthesized and characterized by K.K., A.H.T. and I.P. S.P.G. oversaw the project and edited the manuscript. J.A.P. oversaw the project, performed the experiment for comparison of TMT0 and TMTpro0, ran the MS analysis, performed the data analyses and wrote the manuscript. All authors approved the manuscript. Author contributions |
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| PublicationSubtitle | Techniques for life scientists and chemists |
| PublicationTitle | Nature methods |
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| References | Dikic, Elazar (CR20) 2018; 19 Dai (CR29) 2019; 88 Paulo (CR8) 2015; 15 Savitski, Wilhelm, Hahne, Kuster, Bantscheff (CR38) 2015; 14 Kim, Guan (CR15) 2019; 21 Barilari (CR24) 2017; 36 Harding (CR27) 2018; 46 Huang da, Sherman, Lempicki (CR40) 2009; 4 CR14 Saxton, Sabatini (CR16) 2017; 169 Rauniyar, Yates (CR4) 2014; 13 McAlister (CR12) 2014; 86 Huttlin (CR37) 2010; 143 Elias, Gygi (CR36) 2007; 4 Savitski (CR30) 2018; 173 Chick (CR6) 2016; 534 Navarrete-Perea, Yu, Gygi, Paulo (CR18) 2018; 17 Gaetani (CR25) 2019; 18 Thompson (CR3) 2003; 75 Hebert (CR33) 2013; 10 Mulvey (CR34) 2017; 12 Beausoleil, Villen, Gerber, Rush, Gygi (CR39) 2006; 24 Dephoure, Gygi (CR32) 2012; 5 Vasaikar (CR5) 2019; 177 Ben-Sahra, Howell, Asara, Manning (CR23) 2013; 339 Erickson (CR13) 2019; 18 Savitski (CR26) 2014; 346 Becher (CR28) 2016; 12 Pappireddi, Martin, Wuhr (CR2) 2019; 20 Dayon (CR7) 2008; 80 Wu (CR22) 2011; 10 Thoreen (CR17) 2009; 284 Hughes (CR35) 2019; 14 McAlister (CR10) 2012; 84 Zecha (CR31) 2018; 17 Stepanova, Gygi, Paulo (CR9) 2018; 17 Thompson (CR11) 2019; 91 Mizushima (CR19) 2018; 20 An (CR21) 2019; 74 Li (CR1) 2017; 52 M Gaetani (781_CR25) 2019; 18 EL Huttlin (781_CR37) 2010; 143 L Dayon (781_CR7) 2008; 80 BK Erickson (781_CR13) 2019; 18 L Dai (781_CR29) 2019; 88 JA Paulo (781_CR8) 2015; 15 R Wu (781_CR22) 2011; 10 I Ben-Sahra (781_CR23) 2013; 339 J Zecha (781_CR31) 2018; 17 MM Savitski (781_CR26) 2014; 346 CS Hughes (781_CR35) 2019; 14 A Thompson (781_CR11) 2019; 91 SD Harding (781_CR27) 2018; 46 CM Mulvey (781_CR34) 2017; 12 N Mizushima (781_CR19) 2018; 20 W Huang da (781_CR40) 2009; 4 GC McAlister (781_CR12) 2014; 86 MM Savitski (781_CR30) 2018; 173 JM Chick (781_CR6) 2016; 534 GC McAlister (781_CR10) 2012; 84 CC Thoreen (781_CR17) 2009; 284 J Kim (781_CR15) 2019; 21 MM Savitski (781_CR38) 2015; 14 N Rauniyar (781_CR4) 2014; 13 M Barilari (781_CR24) 2017; 36 RA Saxton (781_CR16) 2017; 169 JE Elias (781_CR36) 2007; 4 I Becher (781_CR28) 2016; 12 A Thompson (781_CR3) 2003; 75 781_CR14 SA Beausoleil (781_CR39) 2006; 24 N Dephoure (781_CR32) 2012; 5 J Navarrete-Perea (781_CR18) 2018; 17 N Pappireddi (781_CR2) 2019; 20 S Vasaikar (781_CR5) 2019; 177 H An (781_CR21) 2019; 74 I Dikic (781_CR20) 2018; 19 H Li (781_CR1) 2017; 52 AS Hebert (781_CR33) 2013; 10 E Stepanova (781_CR9) 2018; 17 |
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| Snippet | Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here an expanded set of 16 isobaric reagents based on... |
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| SubjectTerms | 631/1647/2067 631/1647/296 631/45/475 Affinity labeling Bioinformatics Biological Microscopy Biological Techniques Biomedical and Life Sciences Biomedical Engineering/Biotechnology Cell Line Cell lines Humans Isotope Labeling Kinases Labeling Life Sciences Methods Peptides Peptides - chemistry Proline Proteins Proteome - chemistry Proteomes Proteomics Proteomics - methods Reagents Staurosporine Tandem Mass Spectrometry - methods Thermal stability |
| Title | TMTpro reagents: a set of isobaric labeling mass tags enables simultaneous proteome-wide measurements across 16 samples |
| URI | https://link.springer.com/article/10.1038/s41592-020-0781-4 https://www.ncbi.nlm.nih.gov/pubmed/32203386 https://www.proquest.com/docview/2386355443 https://search.proquest.com/docview/2382655747 https://pubmed.ncbi.nlm.nih.gov/PMC7302421 |
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