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Molecular & Cellular Proteomics : MCP Jul 2023Recent advances in mass spectrometry-based proteomics enable the acquisition of increasingly large datasets within relatively short times, which exposes bottlenecks in...
Recent advances in mass spectrometry-based proteomics enable the acquisition of increasingly large datasets within relatively short times, which exposes bottlenecks in the bioinformatics pipeline. Although peptide identification is already scalable, most label-free quantification (LFQ) algorithms scale quadratic or cubic with the sample numbers, which may even preclude the analysis of large-scale data. Here we introduce directLFQ, a ratio-based approach for sample normalization and the calculation of protein intensities. It estimates quantities via aligning samples and ion traces by shifting them on top of each other in logarithmic space. Importantly, directLFQ scales linearly with the number of samples, allowing analyses of large studies to finish in minutes instead of days or months. We quantify 10,000 proteomes in 10 min and 100,000 proteomes in less than 2 h, a 1000-fold faster than some implementations of the popular LFQ algorithm MaxLFQ. In-depth characterization of directLFQ reveals excellent normalization properties and benchmark results, comparing favorably to MaxLFQ for both data-dependent acquisition and data-independent acquisition. In addition, directLFQ provides normalized peptide intensity estimates for peptide-level comparisons. It is an important part of an overall quantitative proteomic pipeline that also needs to include high sensitive statistical analysis leading to proteoform resolution. Available as an open-source Python package and a graphical user interface with a one-click installer, it can be used in the AlphaPept ecosystem as well as downstream of most common computational proteomics pipelines.
Topics: Proteome; Proteomics; Ecosystem; Peptides; Mass Spectrometry; Software
PubMed: 37225017
DOI: 10.1016/j.mcpro.2023.100581 -
Journal of Proteome Research Jan 2021Protein -acylation (commonly known as palmitoylation) is a widespread reversible lipid modification, which plays critical roles in regulating protein localization,... (Review)
Review
Protein -acylation (commonly known as palmitoylation) is a widespread reversible lipid modification, which plays critical roles in regulating protein localization, activity, stability, and complex formation. The deregulation of protein -acylation contributes to many diseases such as cancer and neurodegenerative disorders. The past decade has witnessed substantial progress in proteomic analysis of protein -acylation, which significantly advanced our understanding of -acylation biology. In this review, we summarized the techniques for the enrichment of -acylated proteins or peptides, critically reviewed proteomic studies of protein -acylation at eight different levels, and proposed major challenges for the -acylproteomics field. In summary, proteome-scale analysis of protein -acylation comes of age and will play increasingly important roles in discovering new disease mechanisms, biomarkers, and therapeutic targets.
Topics: Acylation; Lipoylation; Protein S; Proteome; Proteomics
PubMed: 33253586
DOI: 10.1021/acs.jproteome.0c00409 -
Current Opinion in Chemical Biology Feb 2022Tight regulation of protein translation drives the proteome to undergo changes under influence of extracellular or intracellular signals. Despite mass spectrometry-based... (Review)
Review
Tight regulation of protein translation drives the proteome to undergo changes under influence of extracellular or intracellular signals. Despite mass spectrometry-based proteomics being an excellent method to study differences in protein abundance in complex proteomes, analyzing minute or rapid changes in protein synthesis and abundance remains challenging. Therefore, several dedicated techniques to directly detect and quantify newly synthesized proteins have been developed, notably puromycin-based, bio-orthogonal noncanonical amino acid tagging-based, and stable isotope labeling by amino acids in cell culture-based methods, combined with mass spectrometry. These techniques have enabled the investigation of perturbations, stress, or stimuli on protein synthesis. Improvements of these methods are still necessary to overcome various remaining limitations. Recent improvements include enhanced enrichment approaches and combinations with various stable isotope labeling techniques, which allow for more accurate analysis and comparison between conditions on shorter timeframes and in more challenging systems. Here, we aim to review the current state in this field.
Topics: Amino Acids; Isotope Labeling; Mass Spectrometry; Proteome; Proteomics
PubMed: 34364788
DOI: 10.1016/j.cbpa.2021.07.001 -
Expert Review of Proteomics Feb 2021The goal of this review is to highlight the triumphs and frontiers in measurement of the lens proteome as it relates to onset of age-related nuclear cataract. As global... (Review)
Review
INTRODUCTION
The goal of this review is to highlight the triumphs and frontiers in measurement of the lens proteome as it relates to onset of age-related nuclear cataract. As global life expectancy increases, so too does the frequency of age-related nuclear cataracts. Molecular therapeutics do not exist for delay or relief of cataract onset in humans. Since lens fiber cells are incapable of protein synthesis after initial maturation, age-related changes in proteome composition and post-translational modification accumulation can be measured with various techniques. Several of these modifications have been associated with cataract onset.
AREAS COVERED
We discuss the impact of long-lived proteins on the lens proteome and lens homeostasis as well as proteomic techniques that may be used to measure proteomes at various levels of proteomic specificity and spatial resolution.
EXPERT OPINION
There is clear evidence that several proteome modifications are correlated with cataract formation. Past studies should be enhanced with cutting-edge, spatially resolved mass spectrometry techniques to enhance the specificity and sensitivity of modification detection as it relates to cataract formation.
Topics: Cataract; Humans; Lens, Crystalline; Mass Spectrometry; Proteome; Proteomics
PubMed: 33849365
DOI: 10.1080/14789450.2021.1913062 -
Current Topics in Microbiology and... 2019Microorganisms living in community are critical to life on Earth, playing numerous and profound roles in the environment and human and animal health. Though their... (Review)
Review
Microorganisms living in community are critical to life on Earth, playing numerous and profound roles in the environment and human and animal health. Though their essentiality to life is clear, the mechanistic underpinnings of community structure, interactions, and functions are largely unexplored and in need of function-dependent technologies to unravel the mysteries. Activity-based protein profiling offers unprecedented molecular-level characterization of functions within microbial communities and provides an avenue to determine how external exposures result in functional alterations to microbiomes. Herein, we illuminate the current state and prospective contributions of ABPP as it relates to microbial communities. We provide details on the design, development, and validation of probes, challenges associated with probing in complex microbial communities, provide some specific examples of the biological applications of ABPP in microbes and microbial communities, and highlight potential areas for development. The future of ABPP holds real promise for understanding and considerable impact in microbiome studies associated with personalized medicine, precision agriculture, veterinary health, environmental studies, and beyond.
Topics: Animals; Humans; Microbiological Techniques; Microbiota; Proteome; Proteomics
PubMed: 30406866
DOI: 10.1007/82_2018_128 -
Journal of Proteome Research Mar 2018The Consortium for Top-Down Proteomics (CTDP) proposes a standardized notation, ProForma, for writing the sequence of fully characterized proteoforms. ProForma provides...
The Consortium for Top-Down Proteomics (CTDP) proposes a standardized notation, ProForma, for writing the sequence of fully characterized proteoforms. ProForma provides a means to communicate any proteoform by writing the amino acid sequence using standard one-letter notation and specifying modifications or unidentified mass shifts within brackets following certain amino acids. The notation is unambiguous, human-readable, and can easily be parsed and written by bioinformatic tools. This system uses seven rules and supports a wide range of possible use cases, ensuring compatibility and reproducibility of proteoform annotations. Standardizing proteoform sequences will simplify storage, comparison, and reanalysis of proteomic studies, and the Consortium welcomes input and contributions from the research community on the continued design and maintenance of this standard.
Topics: Amino Acid Sequence; Computational Biology; Databases, Protein; Humans; Information Dissemination; International Cooperation; Molecular Sequence Annotation; Protein Processing, Post-Translational; Proteome; Proteomics; Reproducibility of Results; Software; Tandem Mass Spectrometry
PubMed: 29397739
DOI: 10.1021/acs.jproteome.7b00851 -
The Journal of Investigative Dermatology Aug 2023Hand eczema (HE) is a prevalent skin disease. However, the classification of HE into different subtypes remains challenging. A limited number of previous studies have...
Hand eczema (HE) is a prevalent skin disease. However, the classification of HE into different subtypes remains challenging. A limited number of previous studies have employed invasive biopsy-based strategies; yet, studies of the HE proteome using noninvasive tape-stripping methodology have not been reported. In this study, we wanted to assess whether global proteomic analysis of skin tape strip samples can be used for subclassification of patients with HE. Tape strips were collected from patients with HE and healthy skin. Liquid chromatography-mass spectrometry proteomics was performed, and the global protein expression was analyzed. We identified 2,919 proteins in stratum corneum-derived skin cells from tape strip samples. Compared with healthy skin, the lesional samples from patients with HE exhibited increased expression of immune-related markers and a decreased expression of structural barrier proteins. The difference between HE subtypes was restricted to the lesional skin areas and included an increased expression of skin barrier-related proteins independently of the concurrent AD. In conclusion, we found that the noninvasive tape strip method used in combination with liquid chromatography-mass spectrometry proteomics can be used for analysis of skin protein expression in patients with HE. Thus, the method shows potential for assessing the proteomic differences between subtypes of HE and biomarker discovery.
Topics: Humans; Proteome; Proteomics; Skin; Epidermis; Eczema; Biomarkers
PubMed: 36773646
DOI: 10.1016/j.jid.2022.12.024 -
Cell Systems Oct 2018The increasing throughput and sharing of proteomics mass spectrometry data have now yielded over one-third of a million public mass spectrometry runs. However, these...
The increasing throughput and sharing of proteomics mass spectrometry data have now yielded over one-third of a million public mass spectrometry runs. However, these discoveries are not continuously aggregated in an open and error-controlled manner, which limits their utility. To facilitate the reusability of these data, we built the MassIVE Knowledge Base (MassIVE-KB), a community-wide, continuously updating knowledge base that aggregates proteomics mass spectrometry discoveries into an open reusable format with full provenance information for community scrutiny. Reusing >31 TB of public human data stored in a mass spectrometry interactive virtual environment (MassIVE), the MassIVE-KB contains >2.1 million precursors from 19,610 proteins (48% larger than before; 97% of the total) and doubles proteome coverage to 6 million amino acids (54% of the proteome) with strict library-scale false discovery controls, thereby providing evidence for 430 proteins for which sufficient protein-level evidence was previously missing. Furthermore, MassIVE-KB can inform experimental design, helps identify and quantify new data, and provides tools for community construction of specialized spectral libraries.
Topics: Algorithms; Biological Variation, Population; Databases, Protein; Humans; Mass Spectrometry; Proteome; Proteomics
PubMed: 30172843
DOI: 10.1016/j.cels.2018.08.004 -
Science China. Life Sciences Jun 2011Viruses replicate and proliferate in host cells while continuously adjusting to and modulating the host environment. They encode a wide spectrum of multifunctional... (Review)
Review
Viruses replicate and proliferate in host cells while continuously adjusting to and modulating the host environment. They encode a wide spectrum of multifunctional proteins, which interplay with and modify proteins in host cells. Viral genomes were chronologically the first to be sequenced. However, the corresponding viral proteomes, the alterations of host proteomes upon viral infection, and the dynamic nature of proteins, such as post-translational modifications, enzymatic cleavage, and activation or destruction by proteolysis, remain largely unknown. Emerging high-throughput techniques, in particular quantitative or semi-quantitative mass spectrometry-based proteomics analysis of viral and cellular proteomes, have been applied to define viruses and their interactions with their hosts. Here, we review the major areas of viral proteomics, including virion proteomics, structural proteomics, viral protein interactomics, and changes to the host cell proteome upon viral infection.
Topics: Animals; Electrophoresis, Gel, Two-Dimensional; Host-Pathogen Interactions; Humans; Isotope Labeling; Mass Spectrometry; Proteome; Proteomics; RNA Interference; RNA Viruses; Viral Proteins; Virion; Virus Diseases; Viruses
PubMed: 21706410
DOI: 10.1007/s11427-011-4177-7 -
Molecular & Cellular Proteomics : MCP Jan 2016Cysteine occupies a unique place in protein chemistry. The nucleophilic thiol group allows cysteine to undergo a broad range of redox modifications beyond classical... (Review)
Review
Cysteine occupies a unique place in protein chemistry. The nucleophilic thiol group allows cysteine to undergo a broad range of redox modifications beyond classical thiol-disulfide redox equilibria, including S-sulfenylation (-SOH), S-sulfinylation (-SO(2)H), S-sulfonylation (-SO(3)H), S-nitrosylation (-SNO), S-sulfhydration (-SSH), S-glutathionylation (-SSG), and others. Emerging evidence suggests that these post-translational modifications (PTM) are important in cellular redox regulation and protection against oxidative damage. Identification of protein targets of thiol redox modifications is crucial to understanding their roles in biology and disease. However, analysis of these highly labile and dynamic modifications poses challenges. Recent advances in the design of probes for thiol redox forms, together with innovative mass spectrometry based chemoproteomics methods make it possible to perform global, site-specific, and quantitative analyses of thiol redox modifications in complex proteomes. Here, we review chemical proteomic strategies used to expand the landscape of thiol redox modifications.
Topics: Animals; Cysteine; Humans; Mass Spectrometry; Oxidation-Reduction; Protein Processing, Post-Translational; Proteome; Proteomics; Reproducibility of Results; Sulfhydryl Compounds
PubMed: 26518762
DOI: 10.1074/mcp.O115.056051