Identification and Characterization of Human Proteoforms by Top-Down LC-21 Tesla FT-ICR Mass Spectrometry
Open accessAbstract
Successful high-throughput characterization of intact proteins from complex biological samples by mass spectrometry requires instrumentation capable of high mass resolving power, mass accuracy, sensitivity, and spectral acquisition rate. These limitations often necessitate the performance of hundreds of LC-MS/MS experiments to obtain reasonable coverage of the targeted proteome, which is still typically limited to molecular weights below 30 kDa. The National High Magnetic Field Laboratory (NHMFL) recently installed a 21 T FT-ICR mass spectrometer, which is part of the NHMFL FT-ICR User Facility and available to all qualified users. Here we demonstrate top-down LC-21 T FT-ICR MS/MS of intact proteins derived from human colorectal cancer cell lysate. We identified a combined total of 684 unique protein entries observed as 3238 unique proteoforms at a 1% false discovery rate, based on rapid, data-dependent acquisition of collision-induced and electron-transfer dissociation tandem mass spectra from just 40 LC-MS/MS experiments. Our identifications included 372 proteoforms with molecular weights over 30 kDa detected at isotopic resolution, which substantially extends the accessible mass range for high-throughput top-down LC-MS/MS.
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The tPTCR strategy heavily outperforms the previous state-of-the-art for TDP of large proteoforms represented by medium/high DDA.
“The described tPTCR strategy heavily outperforms the previous state-of-the-art for TDP of large proteoforms represented by medium/high DDA”
From Proton Transfer Charge Reduction Enables High-Throughput Top-Down Analysis of Large Proteoforms (2019) · cites this paper as [46]
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Cites
The control of the false discovery rate in multiple testing under dependency
Benjamini, Y., & Yekutieli, D. (2001) · The Annals of Statistics · doi:10.1214/aos/1013699998 · Cited by 10,490 · see citing works
Statistical significance for genomewide studies
Storey, J. D., & Tibshirani, R. (2003) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1530509100 · Cited by 9,692 · see citing works
Mass spectrometric sequencing of proteins silver-stained polyacrylamide gels.
Shevchenko, A., Wilm, M., Vorm, O., & Mann, M. F. (1996) · Analytical Chemistry · doi:10.1021/ac950914h · Cited by 9,047 · see citing works
Wessel, D., & Flügge, U. (1984) · Analytical Biochemistry · doi:10.1016/0003-2697(84)90782-6 · Cited by 3,997 · see citing works
Peptide and protein sequence analysis by electron transfer dissociation mass spectrometry
Syka, J. E. P., Coon, J. J., Schroeder, M. J., Shabanowitz, J., & Hunt, D. F. (2004) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.0402700101 · Cited by 2,370 · see citing works
Fourier transform ion cyclotron resonance mass spectrometry: A primer
Marshall, A. G., Hendrickson, C. L., & Jackson, G. S. (1998) · Mass Spectrometry Reviews · doi:10.1002/(sici)1098-2787(1998)17:1 · Cited by 1,992 · see citing works
Self-citationMobile and localized protons: a framework for understanding peptide dissociation
Wysocki, V. H., Tsaprailis, G., Smith, L. L., & Breci, L. A. (2000) · Journal of Mass Spectrometry · doi:10.1002/1096-9888(200012)35:12 · Cited by 998 · see citing works
A two-dimensional quadrupole ion trap mass spectrometer
Schwartz, J. C., Senko, M. W., & Syka, J. E. P. (2002) · Journal of the American Society for Mass Spectrometry · doi:10.1016/s1044-0305(02)00384-7 · Cited by 800 · see citing works
Mapping intact protein isoforms in discovery mode using top-down proteomics
Tran, J. C., Zamdborg, L., Ahlf, D. R., Lee, J. E., Catherman, A. D., Durbin, K. R., Tipton, J. D., Vellaichamy, A., Kellie, J. F., Li, M., et al. (2011) · Nature · doi:10.1038/nature10575 · Cited by 665 · see citing works
Self-citationThe utility of ETD mass spectrometry in proteomic analysis
Mikesh, L. M., Ueberheide, B. M., Chi, A., Coon, J. J., Syka, J. E. P., Shabanowitz, J., & Hunt, D. F. (2006) · Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics · doi:10.1016/j.bbapap.2006.10.003 · Cited by 535 · see citing works
Beckler, M. D., Higginbotham, J. N., Franklin, J. L., Ham, A. L., Halvey, P. J., Imasuen, I., Whitwell, C. A., Li, M., Liebler, D. C., & Coffey, R. J. (2012) · Molecular & Cellular Proteomics · doi:10.1074/mcp.m112.022806 · Cited by 514 · see citing works
Protein identification using sequential ion/ion reactions and tandem mass spectrometry
Coon, J. J., Ueberheide, B., Syka, J. E. P., Dryhurst, D. D., Ausio, J., Shabanowitz, J., & Hunt, D. F. (2005) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.0503189102 · Cited by 373 · see citing works
Complete Protein Characterization Using Top-Down Mass Spectrometry and Ultraviolet Photodissociation
Shaw, J. B., Li, W., Holden, D. D., Zhang, Y., Griep-Raming, J., Fellers, R. T., Early, B. P., Thomas, P. M., Kelleher, N. L., & Brodbelt, J. S. (2013) · Journal of the American Chemical Society · doi:10.1021/ja4029654 · Cited by 371 · see citing works
Self-citationHendrickson, C. L., Quinn, J. P., Kaiser, N. K., Smith, D. F., Blakney, G. T., Chen, T., Marshall, A. G., Weisbrod, C. R., & Beu, S. C. (2015) · Journal of the American Society for Mass Spectrometry · doi:10.1007/s13361-015-1182-2 · Cited by 278 · see citing works
Self-citationFinding one's way in proteomics: a protein species nomenclature
Schlüter, H., Apweiler, R., Holzhütter, H., & Jungblut, P. (2009) · Chemistry Central Journal · doi:10.1186/1752-153x-3-11 · Cited by 254 · see citing works
Zamdborg, L., LeDuc, R. D., Glowacz, K. J., Kim, Y.-B., Viswanathan, V., Spaulding, I. T., Early, B. P., Bluhm, E. J., Babai, S., & Kelleher, N. L. (2007) · Nucleic Acids Research · doi:10.1093/nar/gkm371 · Cited by 251 · see citing works
Self-citationOn the Scalability and Requirements of Whole Protein Mass Spectrometry
Compton, P. D., Zamdborg, L., Thomas, P. M., & Kelleher, N. L. (2011) · Analytical Chemistry · doi:10.1021/ac2010795 · Cited by 243 · see citing works
Self-citationTran, J. C., & Doucette, A. A. (2008) · Analytical Chemistry · doi:10.1021/ac702197w · Cited by 206 · see citing works
High-performance mass spectrometry: Fourier transform ion cyclotron resonance at 14.5 Tesla.
Schaub, T. M., Hendrickson, C., Horning, S., Quinn, J. P., Senko, M., & Marshall, A. (2008) · Analytical Chemistry · doi:10.1021/ac800386h · Cited by 178 · see citing works
Self-citationIon isolation and sequential stages of mass spectrometry in a quadrupole ion trap mass spectrometer
Louris, J. N., Brodbelt-Lustig, J. S., Cooks, R. G., et al. (1990) · International Journal of Mass Spectrometry and Ion Processes · doi:10.1016/0168-1176(90)87025-c · Cited by 167 · see citing works
Catherman, A. D., Durbin, K. R., Ahlf, D. R., Early, B. P., Fellers, R. T., Tran, J. C., Thomas, P. M., & Kelleher, N. L. (2013) · Molecular & Cellular Proteomics · doi:10.1074/mcp.m113.030114 · Cited by 157 · see citing works
Self-citationAnsong, C., Wu, S., Meng, D., Liu, X., Brewer, H. M., Deatherage Kaiser, B. L., Nakayasu, E. S., Cort, J. R., Pevzner, P., Smith, R. D., et al. (2013) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1221210110 · Cited by 157 · see citing works
Reid, G. E., Wu, J., Chrisman, P. A., Wells, J. M., & McLuckey, S. A. (2001) · Analytical Chemistry · doi:10.1021/ac0101095 · Cited by 152 · see citing works
Collisions or Electrons? Protein Sequence Analysis in the 21st Century
Coon, J. J. (2009) · Analytical Chemistry · doi:10.1021/ac802330b · Cited by 149 · see citing works
Top-down mass spectrometry: Recent developments, applications and perspectives
Cui, W., Rohrs, H. W., & Gross, M. L. (2011) · The Analyst · doi:10.1039/c1an15286f · Cited by 140 · see citing works
Quantitation and Identification of Thousands of Human Proteoforms below 30 kDa
Durbin, K. R., Fornelli, L., Fellers, R. T., Doubleday, P. F., Narita, M., & Kelleher, N. L. (2016) · Journal of Proteome Research · doi:10.1021/acs.jproteome.5b00997 · Cited by 111 · see citing works
Self-citationNielsen, M. L., Savitski, M. M., & Zubarev, R. A. (2006) · Molecular & Cellular Proteomics · doi:10.1074/mcp.m600248-mcp200 · Cited by 103 · see citing works
The first pilot project of the consortium for top‐down proteomics: A status report
Dang, X., Scotcher, J., Wu, S., Chu, R., Tolić, N., Ntai, I., Thomas, P. M., Fellers, R. T., Early, B. P., Zheng, Y., et al. (2014) · PROTEOMICS · doi:10.1002/pmic.201300438 · Cited by 101 · see citing works
Self-citationUdeshi, N. D., Compton, P. D., Shabanowitz, J., Hunt, D. F., & Rose, K. L. (2008) · Nature Protocols · doi:10.1038/nprot.2008.159 · Cited by 101 · see citing works
Ultraviolet Photodissociation for Characterization of Whole Proteins on a Chromatographic Time Scale
Cannon, J. R., Cammarata, M. B., Robotham, S. A., Cotham, V. C., Shaw, J., Fellers, R. T., Early, B. P., Thomas, P. M., Kelleher, N. L., & Brodbelt, J. S. (2014) · Analytical Chemistry · doi:10.1021/ac403859a · Cited by 92 · see citing works
Self-citationEvaluation of the Compact High-Field Orbitrap for Top-Down Proteomics of Human Cells
Ahlf, D. R., Compton, P. D., Tran, J. C., Early, B. P., Thomas, P. M., & Kelleher, N. L. (2012) · Journal of Proteome Research · doi:10.1021/pr3004216 · Cited by 89 · see citing works
Self-citationLeDuc, R. D., Fellers, R. T., Early, B. P., Greer, J. B., Thomas, P. M., & Kelleher, N. L. (2014) · Journal of Proteome Research · doi:10.1021/pr401277r · Cited by 85 · see citing works
Self-citationXiu, L., Valeja, S. G., Alpert, A. J., Jin, S., & Ge, Y. (2014) · Analytical Chemistry · doi:10.1021/ac501836k · Cited by 68 · see citing works
Front-End Electron Transfer Dissociation: A New Ionization Source
Earley, L., Anderson, L. C., Bai, D. L., Mullen, C., Syka, J. E. P., English, A. M., Dunyach, J., Stafford, G. C., Shabanowitz, J., Hunt, D. F., et al. (2013) · Analytical Chemistry · doi:10.1021/ac401783f · Cited by 67 · see citing works
Self-citationAnderson, L. C., Karch, K. R., Ugrin, S. A., Coradin, M., English, A. M., Sidoli, S., Shabanowitz, J., Garcia, B. A., & Hunt, D. F. (2016) · Molecular & Cellular Proteomics · doi:10.1074/mcp.o115.053843 · Cited by 60 · see citing works
Self-citationEnhanced Dissociation of Intact Proteins with High Capacity Electron Transfer Dissociation
Riley, N. M., Mullen, C., Weisbrod, C. R., Sharma, S., Senko, M. W., Zabrouskov, V., Westphall, M. S., Syka, J. E. P., & Coon, J. J. (2015) · Journal of the American Society for Mass Spectrometry · doi:10.1007/s13361-015-1306-8 · Cited by 59 · see citing works
Halvey, P. J., Wang, X., Wang, J., et al. (2013) · Cancer Research · doi:10.1158/0008-5472.can-13-2488 · Cited by 49 · see citing works
Controlled Ion Ejection from an External Trap for Extended m/z Range in FT-ICR Mass Spectrometry
Kaiser, N. K., Savory, J., & Hendrickson, C. L. (2014) · Journal of the American Society for Mass Spectrometry · doi:10.1007/s13361-014-0871-6 · Cited by 46 · see citing works
Self-citationScherperel, G., & Reid, G. E. (2007) · The Analyst · doi:10.1039/b618499p · Cited by 30 · see citing works
Savaryn, J. P., Toby, T. K., Catherman, A. D., Fellers, R. T., LeDuc, R. D., Thomas, P. M., Friedewald, J., Salomon, D., Abecassis, M., & Kelleher, N. L. (2016) · Proteomics · doi:10.1002/pmic.201600008 · Cited by 23 · see citing works
Self-citationMeta-analysis for Protein Identification: A Case Study on Yeast Data
Higdon, R., Haynes, W. A., & Kolker, E. (2010) · OMICS A Journal of Integrative Biology · doi:10.1089/omi.2010.0034 · Cited by 16 · see citing works
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Kafader, J. O., Melani, R. D., Durbin, K. R., Ikwuagwu, B., Early, B. P., Fellers, R. T., Beu, S. C., Zabrouskov, V., Makarov, A. A., Maze, J. T., et al. (2020) · Nature Methods · doi:10.1038/s41592-020-0764-5 · Cited by 197 · see citing works
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Macias, L. A., Santos, I. C., & Brodbelt, J. S. (2020) · Analytical Chemistry · doi:10.1021/acs.analchem.9b04859 · Cited by 107 · see citing works
Ntai, I., Fornelli, L., DeHart, C. J., Hutton, J. E., Doubleday, P. F., LeDuc, R. D., van Nispen, A. J., Fellers, R. T., Whiteley, G., Boja, E. S., et al. (2018) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1716122115 · Cited by 108 · see citing works
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- Authors
- Anderson, L. C., DeHart, C. J., Kaiser, N. K., Fellers, R. T., Smith, D. F., Greer, J. B., LeDuc, R. D., Blakney, G. T., Thomas, P. M., Kelleher, N. L., & Hendrickson, C. L.
- Affiliations
- National High Magnetic Field Laboratory (US)
- Northwestern University (US)
- Florida State University (US)
- Corresponding author
- Anderson, Lissa C.
- ORCID
- Venue
- Journal of Proteome Research
- Publisher
- American Chemical Society (ACS)
- Published
- 2016-12-12
- Volume
- 16
- Issue
- 2
- Pages
- 1087-1096
- ISSN
- 1535-3893 (ISSN-L)
- 1535-3907
- PMC
- PMC5353847
- OpenAlex
- W2482415955
- Cited by
- 94
- References
- 41
- Citations by year
- 2026: 3 · 2025: 4 · 2024: 2 · 2023: 3 · 2022: 13 · 2021: 7 · 2020: 15 · 2019: 22 · 2018: 13 · 2017: 11 · 2015: 1
- FWCI
- 7.62
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Journal of Proteome Research (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- IUScholarWorks (Indiana University) (submittedVersion, cc-by-nc)
- Funders
- U.S. Department of Health and Human Services (P41GM108569)
- National Science Foundation (DMR-1157490)
- Topics
- Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
- Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Metabolomics and Mass Spectrometry Studies
- Keywords
- top-down proteomics, proteoforms, high mass resolution, collision-induced dissociation, electron transfer dissociation
- MeSH terms
- Amino Acid Sequence, Fourier Analysis, Humans, Neoplasm Proteins, Mass Spectrometry, Colorectal Neoplasms, Cyclotrons, Proteome, Proteomics, Complex Mixtures
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