2021
DOI: 10.1021/acs.jproteome.1c00357
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Toward Comprehensive Plasma Proteomics by Orthogonal Protease Digestion

Abstract: Rapid and consistent protein identification across large clinical cohorts is an important goal for clinical proteomics. With the development of data-independent technologies (DIA/SWATH-MS), it is now possible to analyze hundreds of samples with great reproducibility and quantitative accuracy. However, this technology benefits from empirically derived spectral libraries that define the detectable set of peptides and proteins. Here, we apply a simple and accessible tip-based workflow for the generation of spectr… Show more

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Cited by 18 publications
(17 citation statements)
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“…Glu-C has also been heavily utilized for plasma proteomic applications. 39,48 There are also proteases which cleave N-terminally, or before their triggering amino acids. Asp-N cleaves before aspartic acid residues.…”
Section: The Value Of Alternative Proteasesmentioning
confidence: 99%
“…Glu-C has also been heavily utilized for plasma proteomic applications. 39,48 There are also proteases which cleave N-terminally, or before their triggering amino acids. Asp-N cleaves before aspartic acid residues.…”
Section: The Value Of Alternative Proteasesmentioning
confidence: 99%
“…The SEC samples were prepared as we previously reported [51] A final fraction at 80% ACN was added to recover hydrophobic peptides.…”
Section: Sec-ms Proteomics Sample Preparationmentioning
confidence: 99%
“…While deconvolving peptide signals from this type of data can be more challenging, the use of peptide-centric searching has significantly improved peptide detection rates. This new paradigm presents the opportunity to multiplex proteomic samples generated from a variety of different proteases in a single DIA-MS analysis, , and consequently significantly increase proteome sequence coverage while incurring minimal increases in MS instrument time. Furthermore, results by Bruderer et al demonstrate the ability to detect more peptides in a single LC-MS/MS experiment than the number of acquired MS/MS spectra, and imply a hidden capacitance of DIA to detect more peptides within a given dynamic range .…”
Section: Introductionmentioning
confidence: 99%
“…Furthermore, results by Bruderer et al demonstrate the ability to detect more peptides in a single LC-MS/MS experiment than the number of acquired MS/MS spectra, and imply a hidden capacitance of DIA to detect more peptides within a given dynamic range . However, to date DIA has been highly optimized for tryptic peptides fragmented by higher energy beam-type CID (bt-CID), with few reported examples of the use of nontryptic proteases , or lower energy resonance excitation CID (re-CID). , As a result, high resolution peptide libraries suitable for analyzing DIA measurements of nontryptic peptides are extremely limited, and methods to generate DIA-only libraries are constrained by lower-accuracy fragmentation prediction for nontryptic peptides using current modeling algorithms …”
Section: Introductionmentioning
confidence: 99%