Abstract
Subfemtomole peptide sequence analysis has been achieved using microcapillary HPLC columns, with integrated nanoelectrospray emitters, coupled directly to a Fourier transform ion cyclotron resonance mass spectrometer. Accurate mass (+/-0.010 Da) peptide maps are generated from a standard six-protein digest mixture, whose principle components span a concentration dynamic range of 1000:1. Iterative searches against approximately 189000 entries in the OWL database readily identify each protein, with high sequence coverage (20-60%), from as little as 10 amol loaded on-column. In addition, a simple variable-flow HPLC apparatus provides for on-line tandem mass spectrometric analysis of tryptic peptides at the 400-amol level. MS/MS data are searched against approximately 280000 entries in a nonredundant protein database using SEQUEST. Accurate precursor and product ion mass information readily identifies primary amino acid sequences differing by asparagine vs aspartic acid (deltam = 0.98 Da) and glutamine vs lysine (deltam = 0.036 Da).
Keywords
ChemistryFourier transform ion cyclotron resonanceMass spectrometryChromatographyTandem mass spectrometryPeptideProtein mass spectrometryIon cyclotron resonanceAnalytical Chemistry (journal)Top-down proteomicsHigh-performance liquid chromatographyPeptide sequenceAsparagineAmino acidIonCyclotronBiochemistryOrganic chemistry
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Publication Info
- Year
- 2000
- Type
- article
- Volume
- 72
- Issue
- 18
- Pages
- 4266-4274
- Citations
- 277
- Access
- Closed
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Cite This
Susan E. Martin,
Jeffrey Shabanowitz,
Donald F. Hunt
et al.
(2000).
Subfemtomole MS and MS/MS Peptide Sequence Analysis Using Nano-HPLC Micro-ESI Fourier Transform Ion Cyclotron Resonance Mass Spectrometry.
Analytical Chemistry
, 72
(18)
, 4266-4274.
https://doi.org/10.1021/ac000497v
Identifiers
- DOI
- 10.1021/ac000497v