Journal
JOURNAL OF PROTEOME RESEARCH
Volume 16, Issue 2, Pages 1087-1096Publisher
AMER CHEMICAL SOC
DOI: 10.1021/acs.jproteome.6b00696
Keywords
FT-ICR; 21 tesla; top-down proteomics; FTMS
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Funding
- National Science Foundation [DMR-1157490]
- State of Florida
- National Institute of General Medical Science [P41GM108569]
- Northwestern University Information Technology
- Office of the Provost
- Office for Research
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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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