Ultra‐high sensitivity mass spectrometry quantifies single‐cell proteome changes upon perturbation

Molecular Systems Biology, Год журнала: 2022, Номер 18(3)

Опубликована: Фев. 28, 2022

Язык: Английский

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dia-PASEF data analysis using FragPipe and DIA-NN for deep proteomics of low sample amounts

Nature Communications, Год журнала: 2022, Номер 13(1)

Опубликована: Июль 8, 2022

The dia-PASEF technology uses ion mobility separation to reduce signal interferences and increase sensitivity in proteomic experiments. Here we present a two-dimensional peak-picking algorithm generation of optimized spectral libraries, as well take advantage neural network-based processing data. Our computational platform boosts depth by up 83% compared previous work, is specifically beneficial for fast experiments those with low sample amounts. It quantifies over 5300 proteins single injections recorded at 200 samples per day throughput using Evosep One chromatography system on timsTOF Pro mass spectrometer almost 9000 93-min nanoflow gradient 2, from ng HeLa peptides. A user-friendly implementation provided through the incorporation algorithms DIA-NN software FragPipe workflow library generation.

Язык: Английский

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Increasing the throughput of sensitive proteomics by plexDIA

Nature Biotechnology, Год журнала: 2022, Номер 41(1), С. 50 - 59

Опубликована: Июль 14, 2022

Язык: Английский

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Advances and Utility of the Human Plasma Proteome

Journal of Proteome Research, Год журнала: 2021, Номер 20(12), С. 5241 - 5263

Опубликована: Окт. 21, 2021

The study of proteins circulating in blood offers tremendous opportunities to diagnose, stratify, or possibly prevent diseases. With recent technological advances and the urgent need understand effects COVID-19, proteomic analysis blood-derived serum plasma has become even more important for studying human biology pathophysiology. Here we provide views perspectives about developments possible clinical applications that use mass-spectrometry(MS)- affinity-based methods. We discuss examples where proteomics contributed valuable insights into SARS-CoV-2 infections, aging, hemostasis offered by combining with genetic data. As a contribution Human Proteome Organization (HUPO) Plasma Project (HPPP), present PeptideAtlas build 2021-07 comprises 4395 canonical 1482 additional nonredundant detected 240 MS-based experiments. In addition, report new Extracellular Vesicle 2021-06, which five studies 2757 extracellular vesicles blood, 74% (2047) are common PeptideAtlas. Our overview summarizes advances, impactful applications, ongoing challenges translating utility precision medicine.

Язык: Английский

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Trapped Ion Mobility Spectrometry and Parallel Accumulation–Serial Fragmentation in Proteomics

Molecular & Cellular Proteomics, Год журнала: 2021, Номер 20, С. 100138 - 100138

Опубликована: Янв. 1, 2021

Recent advances in efficiency and ease of implementation have rekindled interest ion mobility spectrometry, a technique that separates gas phase ions by their size shape can be hybridized with conventional LC MS. Here, we review the recent development trapped spectrometry (TIMS) coupled to TOF mass analysis. In particular, parallel accumulation–serial fragmentation (PASEF) operation mode offers unique advantages terms sequencing speed sensitivity. Its defining feature is it synchronizes release from TIMS device downstream selection precursors for quadrupole configuration. As are compressed into narrow peaks, number peptide fragment spectra obtained data-dependent or targeted analyses increased an order magnitude without compromising Taking advantage correlation between mass, PASEF principle also multiplies data-independent acquisition. This makes technology well suited rapid proteome profiling, increasingly important attribute clinical proteomics, as ultrasensitive measurements down single cells. The accuracy enable precise collisional cross section values at scale more than million data points neural networks capable predicting them based only on sequences. Peptide differ isobaric sequences positional isomers post-translational modifications. additional information may leveraged real time direct acquisition postprocessing increase confidence identifications. These developments make powerful expandable platform proteomics beyond.

Язык: Английский

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Ultra-fast label-free quantification and comprehensive proteome coverage with narrow-window data-independent acquisition

Nature Biotechnology, Год журнала: 2024, Номер unknown

Опубликована: Фев. 1, 2024

Abstract Mass spectrometry (MS)-based proteomics aims to characterize comprehensive proteomes in a fast and reproducible manner. Here we present the narrow-window data-independent acquisition (nDIA) strategy consisting of high-resolution MS1 scans with parallel tandem MS (MS/MS) ~200 Hz using 2-Th isolation windows, dissolving differences between data-dependent -independent methods. This is achieved by pairing quadrupole Orbitrap mass spectrometer asymmetric track lossless (Astral) analyzer which provides >200-Hz MS/MS scanning speed, high resolving power sensitivity, low-ppm accuracy. The nDIA enables profiling >100 full yeast per day, or 48 human day at depth ~10,000 protein groups half-an-hour ~7,000 proteins 5 min, representing 3× higher coverage compared current state-of-the-art MS. Multi-shot offline fractionated samples ~3 h. High quantitative precision accuracy are demonstrated three-species proteome mixture, quantifying 14,000+ single run.

Язык: Английский

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MSBooster: improving peptide identification rates using deep learning-based features

Nature Communications, Год журнала: 2023, Номер 14(1)

Опубликована: Июль 27, 2023

Peptide identification in liquid chromatography-tandem mass spectrometry (LC-MS/MS) experiments relies on computational algorithms for matching acquired MS/MS spectra against sequences of candidate peptides using database search tools, such as MSFragger. Here, we present a new tool, MSBooster, rescoring peptide-to-spectrum matches additional features incorporating deep learning-based predictions peptide properties, LC retention time, ion mobility, and spectra. We demonstrate the utility tandem with MSFragger Percolator, several different workflows, including nonspecific searches (immunopeptidomics), direct from data independent acquisition data, single-cell proteomics, generated an mobility separation-enabled timsTOF MS platform. MSBooster is fast, robust, fully integrated into widely used FragPipe

Язык: Английский

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Advances in data‐independent acquisition mass spectrometry towards comprehensive digital proteome landscape

Mass Spectrometry Reviews, Год журнала: 2022, Номер 42(6), С. 2324 - 2348

Опубликована: Май 29, 2022

Abstract The data‐independent acquisition mass spectrometry (DIA‐MS) has rapidly evolved as a powerful alternative for highly reproducible proteome profiling with unique strength of generating permanent digital maps retrospective analysis biological systems. Recent advancements in data software tools the complex DIA‐MS/MS spectra coupled to fast MS scanning speed and high accuracy have greatly expanded sensitivity coverage DIA‐based proteomics profiling. Here, we review evolution DIA‐MS techniques, from earlier proof‐of‐principle parallel fragmentation all‐ions or ions selected m / z range, sequential window all theoretical (SWATH‐MS) latest innovations, recent development computation algorithms informatics, auxiliary advanced instrumentation enhance performance DIA‐MS. We further summarize applications experimentally‐derived well silico library resources large‐scale facilitate biomarker discovery drug human diseases emphasis on proteomic coverage. Toward next‐generation clinical proteomics, outline challenges processing multi‐dimensional DIA set continuing need higher sensitivity.

Язык: Английский

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Microbial communities form rich extracellular metabolomes that foster metabolic interactions and promote drug tolerance

Nature Microbiology, Год журнала: 2022, Номер 7(4), С. 542 - 555

Опубликована: Март 21, 2022

Abstract Microbial communities are composed of cells varying metabolic capacity, and regularly include auxotrophs that lack essential pathways. Through analysis for amino acid biosynthesis pathways in microbiome data derived from >12,000 natural microbial obtained as part the Earth Microbiome Project (EMP), study auxotrophic–prototrophic interactions self-establishing metabolically cooperating yeast (SeMeCos), we reveal a imprinted mechanism links presence to an increase gains antimicrobial drug tolerance. As consequence adaptations necessary uptake specific metabolites, obtain altered flux distributions, export more metabolites and, this way, enrich community environments metabolites. Moreover, increased efflux activities reduce intracellular concentrations, allowing grow levels above minimal inhibitory concentrations. For example, show antifungal action azoles is greatly diminished enriched environment. Our results hence provide explains why robust exposure when they interact metabolically.

Язык: Английский

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MS-Based Proteomics of Body Fluids: The End of the Beginning

Molecular & Cellular Proteomics, Год журнала: 2023, Номер 22(7), С. 100577 - 100577

Опубликована: Май 19, 2023

Accurate biomarkers are a crucial and necessary precondition for precision medicine, yet existing ones often unspecific new have been very slow to enter the clinic. Mass spectrometry (MS)-based proteomics excels by its untargeted nature, specificity of identification, quantification, making it an ideal technology biomarker discovery routine measurement. It has unique attributes compared affinity binder technologies, such as OLINK Proximity Extension Assay SOMAscan. In in previous review 2017, we described technological conceptual limitations that had held back success. We proposed 'rectangular strategy' better separate true minimizing cohort-specific effects. Today, this converged with advances MS-based technology, increased sample throughput, depth quantification. As result, studies become more successful, producing candidates withstand independent verification and, some cases, already outperform state-of-the-art clinical assays. summarize developments over last years, including benefits large cohorts, which acceptance. Shorter gradients, scan modes, multiplexing about drastically increase cross-study integration, proxies absolute levels. found multiprotein panels inherently robust than current single analyte tests capture complexity human phenotypes. Routine MS measurement clinic is fast becoming viable option. The full set proteins body fluid (global proteome) most important reference best process control. Additionally, increasingly all information could be obtained from targeted analysis although latter may straightforward way regular use. Many challenges remain, not least regulatory ethical but outlook applications never brighter.

Язык: Английский

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