An Optimized Shotgun Strategy for the Rapid Generation of Comprehensive Human Proteomes
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An Optimized Shotgun Strategy for the Rapid Generation of Comprehensive Human Proteomes. / Bekker-Jensen, Dorte B; Kelstrup, Christian D; Batth, Tanveer S; Larsen, Sara C; Haldrup, Christa; Bramsen, Jesper B; Sørensen, Karina D; Høyer, Søren; Ørntoft, Torben F; Andersen, Claus L; Nielsen, Michael L; Olsen, Jesper V.
In: Cell Systems, Vol. 4, No. 6, 2017, p. 587-599, e1-e4.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - An Optimized Shotgun Strategy for the Rapid Generation of Comprehensive Human Proteomes
AU - Bekker-Jensen, Dorte B
AU - Kelstrup, Christian D
AU - Batth, Tanveer S
AU - Larsen, Sara C
AU - Haldrup, Christa
AU - Bramsen, Jesper B
AU - Sørensen, Karina D
AU - Høyer, Søren
AU - Ørntoft, Torben F
AU - Andersen, Claus L
AU - Nielsen, Michael L
AU - Olsen, Jesper V
N1 - Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.
PY - 2017
Y1 - 2017
N2 - This study investigates the challenge of comprehensively cataloging the complete human proteome from a single-cell type using mass spectrometry (MS)-based shotgun proteomics. We modify a classical two-dimensional high-resolution reversed-phase peptide fractionation scheme and optimize a protocol that provides sufficient peak capacity to saturate the sequencing speed of modern MS instruments. This strategy enables the deepest proteome of a human single-cell type to date, with the HeLa proteome sequenced to a depth of ∼584,000 unique peptide sequences and ∼14,200 protein isoforms (∼12,200 protein-coding genes). This depth is comparable with next-generation RNA sequencing and enables the identification of post-translational modifications, including ∼7,000 N-acetylation sites and ∼10,000 phosphorylation sites, without the need for enrichment. We further demonstrate the general applicability and clinical potential of this proteomics strategy by comprehensively quantifying global proteome expression in several different human cancer cell lines and patient tissue samples.
AB - This study investigates the challenge of comprehensively cataloging the complete human proteome from a single-cell type using mass spectrometry (MS)-based shotgun proteomics. We modify a classical two-dimensional high-resolution reversed-phase peptide fractionation scheme and optimize a protocol that provides sufficient peak capacity to saturate the sequencing speed of modern MS instruments. This strategy enables the deepest proteome of a human single-cell type to date, with the HeLa proteome sequenced to a depth of ∼584,000 unique peptide sequences and ∼14,200 protein isoforms (∼12,200 protein-coding genes). This depth is comparable with next-generation RNA sequencing and enables the identification of post-translational modifications, including ∼7,000 N-acetylation sites and ∼10,000 phosphorylation sites, without the need for enrichment. We further demonstrate the general applicability and clinical potential of this proteomics strategy by comprehensively quantifying global proteome expression in several different human cancer cell lines and patient tissue samples.
KW - Journal Article
U2 - 10.1016/j.cels.2017.05.009
DO - 10.1016/j.cels.2017.05.009
M3 - Journal article
C2 - 28601559
VL - 4
SP - 587-599, e1-e4
JO - Cell Systems
JF - Cell Systems
SN - 2405-4712
IS - 6
ER -
ID: 184291218