PRM-LIVE with Trapped Ion Mobility Spectrometry and Its Application in Selectivity Profiling of Kinase Inhibitors

Zhu, He; Ficarro, Scott B.; Alexander, William M.; Fleming, Laura E; Adelmant, Guillaume; Zhang, Tinghu; Willetts, Matthew; Decker, Jens; Brehmer, Sven; Krause, Michael; East, Michael P.; Gray, Nathanael S.; Johnson, Gary L.; Kruppa, Gary; Marto, Jarrod A.

doi:10.1021/acs.analchem.1c02349

Cited by 23 publications

(33 citation statements)

References 47 publications

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“…1. This tiered approach may also be implemented with other approaches for performing real-time retention time alignment 33, 34 .…”

Section: Discussionmentioning

confidence: 99%

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Prioritized single-cell proteomics reveals molecular and functional polarization across primary macrophages

Huffman¹,

Leduc²,

Wichmann

et al. 2022

Preprint

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Major aims of single-cell proteomics include increasing the consistency, sensitivity, and depth of protein quantification, especially for proteins and modifications of biological interest. To simultaneously advance all of these aims, we developed prioritized Single Cell ProtEomics (pSCoPE). pSCoPE ensures duty-cycle time for analyzing prioritized peptides across all single cells (thus increasing data consistency) while analyzing identifiable peptides at full duty-cycle, thus increasing proteome depth. These strategies increased the quantified data points for challenging peptides and the overall proteome coverage about 2-fold. pSCoPE enabled quantifying proteome polarization in primary mouse macrophages and linking it to phenotypic variability in endocytic activity. Proteins annotated to phagosome maturation and proton transport showed concerted variation for both untreated and lipopolysaccharide-treated macrophages, indicating a conserved axis of polarization. pSCoPE further quantified proteolytic products, suggesting a gradient of cathepsin activities within a treatment condition. pSCoPE is easily accessible and likely to benefit many applications, especially mechanistic analysis seeking to focus on proteins of interest without sacrificing proteome coverage.

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“…1. This tiered approach may also be implemented with other approaches for performing real-time retention time alignment 33, 34 .…”

Section: Discussionmentioning

confidence: 99%

“…This prioritized algorithm (Fig. 1) introduced here may also be implemented with other approaches for performing real-time retention time alignment 44, 45 and may be extended to single-cell proteomics multiplexed by non-isobaric mass tags 46 .…”

Section: Discussionmentioning

confidence: 99%

Prioritized single-cell proteomics reveals molecular and functional polarization across primary macrophages

Huffman¹,

Leduc²,

Wichmann

et al. 2022

Preprint

View full text Add to dashboard Cite

show abstract

“…As such, QCPA assays can be used with both high-resolution ion trap and timeof-flight mass spectrometers, including diverse quantitative targeted methods such as SureQuant [19], AIM [23] and others. In addition, the use of QCPA libraries of isotopically labeled peptides permits isotopically-triggered scanning and other automated mass spectrometry acquisition methods [17,28,47,48] to increase assay multiplexing and throughput [4,49]. While specific methods and applications will require explicit optimization of the analytical performance [7,50], the breadth and depth of QCPA targets should enable increasingly comprehensive and integrated measurements of cell signaling.…”

Section: Discussionmentioning

confidence: 99%

Quantitative Cell Proteomic Atlas: Pathway-scale targeted mass spectrometry for high-resolution functional profiling of cell signaling

Cifani

Kentsis

2022

Preprint

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In spite of extensive studies of cellular signaling, many fundamental processes such as pathway integration, cross-talk and feedback remain poorly understood. To enable integrated and quantitative measurements of cellular biochemical activities, we have developed the Quantitative Cell Proteomics Atlas (QCPA). QCPA consists of panels of targeted mass spectrometry assays to determine the abundance and stoichiometry of regulatory post-translational modifications of sentinel proteins from most known physiologic and pathogenic signaling pathways in human cells. QCPA currently profiles 1,913 peptides from 469 effectors of cell surface signaling, apoptosis, stress response, gene expression, quiescence, and proliferation. For each protein, QCPA includes triplets of isotopically labeled peptides covering known post-translational regulatory sites to determine their stoichiometries and unmodified protein regions to measure total protein abundance. The QCPA framework incorporates analytes to control for technical variability of sample preparation and mass spectrometric analysis, including TrypQuant, a synthetic substrate for accurate quantification of proteolysis efficiency for proteins containing chemically modified residues. The ability to precisely and accurately quantify most known signaling pathways should enable improved chemoproteomic approaches for the comprehensive analysis of cell signaling and clinical proteomics of diagnostic specimens. QCPA is openly available at https://qcpa.mskcc.org.

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“…We noticed that Jarrod A. Marto group has just achieved comparable throughput with PRM-LIVE, an extensible, Python-based acquisition engine for the timsTOF Pro, which dynamically adjusts detection windows for reproducible target scheduling. (Zhu et al, 2021), It implies that our strategy is easy to implement in other laboratories with the support of the Cancer Serum Atlas. Researchers can quickly and efficiently find interested proteins and develop the SPA-PRM method with the information from the library, enables simultaneous biomarker discovery and validation.…”

Section: Discussionmentioning

confidence: 99%

Cancer Serum Atlas supported precise pan-targeted proteomics enable multi-cancer detection

Zhang

Yuan

et al. 2022

Preprint

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The wide dynamic range of serum proteome restrained discovery of the clinically interested proteins in large cohort studies. Herein, we presented a high-sensitivity, high-throughput and precise pan-targeted serum proteomic strategy for high-efficient cancer serum proteomic research and biomarker discovery. We constructed a resource of over 2000 cancer-secreted proteins and the standard MS assays and spectra of at least one synthetic unique peptide per protein were acquired and documented (Cancer Serum Atlas,www.cancerserumatlas.com). Then, the standard peptides anchored parallel reaction monitoring (SPA-PRM) method was developed with support of Cancer Serum Atlas, achieving precise quantification of cancer-secreted proteins with high throughput and sensitivity. We directly quantified 325 cancer-related serum proteins in 288 serum of four cancer types (liver, stomach, lung, breast) and controls with the pan-targeted strategy, and discovered considerable potential biomarkers benefit for early detection of cancer. Finally, a proteomics based multi-cancer detection model was built, demonstrating high sensitivity (87.2%), specificity (100%), with 73.8% localization accuracy for an independent test set. In conclusion, the Cancer Serum Atlas provides a wide range of potential biomarkers that serve as targets and standard assays for systematic and high-efficient serological studies of cancer, and the Cancer Serum Atlas supported pan-targeted proteomic strategy enables high-efficient biomarker discovery and multi-cancer detection, thus can be a powerful tool for liquid biopsy.

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PRM-LIVE with Trapped Ion Mobility Spectrometry and Its Application in Selectivity Profiling of Kinase Inhibitors

Cited by 23 publications

References 47 publications

Prioritized single-cell proteomics reveals molecular and functional polarization across primary macrophages

Prioritized single-cell proteomics reveals molecular and functional polarization across primary macrophages

Quantitative Cell Proteomic Atlas: Pathway-scale targeted mass spectrometry for high-resolution functional profiling of cell signaling

Cancer Serum Atlas supported precise pan-targeted proteomics enable multi-cancer detection

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