| 1. |
- Bichmann, Leon, et al.
(författare)
-
DIAproteomics : A Multifunctional Data Analysis Pipeline for Data-Independent Acquisition Proteomics and Peptidomics
- 2021
-
Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 20:7, s. 3758-3766
-
Tidskriftsartikel (refereegranskat)abstract
- Data-independent acquisition (DIA) is becoming a leading analysis method in biomedical mass spectrometry. The main advantages include greater reproducibility and sensitivity and a greater dynamic range compared with data-dependent acquisition (DDA). However, the data analysis is complex and often requires expert knowledge when dealing with large-scale data sets. Here we present DIAproteomics, a multifunctional, automated, high-throughput pipeline implemented in the Nextflow workflow management system that allows one to easily process proteomics and peptidomics DIA data sets on diverse compute infrastructures. The central components are well-established tools such as the OpenSwathWorkflow for the DIA spectral library search and PyProphet for the false discovery rate assessment. In addition, it provides options to generate spectral libraries from existing DDA data and to carry out the retention time and chromatogram alignment. The output includes annotated tables and diagnostic visualizations from the statistical postprocessing and computation of fold-changes across pairwise conditions, predefined in an experimental design. DIAproteomics is well documented open-source software and is available under a permissive license to the scientific community at https://www. openms.de/diaprotcoinics/.
|
|
| 2. |
- Bludau, Isabell, et al.
(författare)
-
Complex-centric proteome profiling by SEC-SWATH-MS for the parallel detection of hundreds of protein complexes
- 2020
-
Ingår i: Nature Protocols. - : Springer Science and Business Media LLC. - 1754-2189 .- 1750-2799. ; 15:8, s. 2341-2386
-
Tidskriftsartikel (refereegranskat)abstract
- Most catalytic, structural and regulatory functions of the cell are carried out by functional modules, typically complexes containing or consisting of proteins. The composition and abundance of these complexes and the quantitative distribution of specific proteins across different modules are therefore of major significance in basic and translational biology. However, detection and quantification of protein complexes on a proteome-wide scale is technically challenging. We have recently extended the targeted proteomics rationale to the level of native protein complex analysis (complex-centric proteome profiling). The complex-centric workflow described herein consists of size exclusion chromatography (SEC) to fractionate native protein complexes, data-independent acquisition mass spectrometry to precisely quantify the proteins in each SEC fraction based on a set of proteotypic peptides and targeted, complex-centric analysis where prior information from generic protein interaction maps is used to detect and quantify protein complexes with high selectivity and statistical error control via the computational framework CCprofiler (https://github.com/CCprofiler/CCprofiler). Complex-centric proteome profiling captures most proteins in complex-assembled state and reveals their organization into hundreds of complexes and complex variants observable in a given cellular state. The protocol is applicable to cultured cells and can potentially also be adapted to primary tissue and does not require any genetic engineering of the respective sample sources. At present, it requires ~8 d of wet-laboratory work, 15 d of mass spectrometry measurement time and 7 d of computational analysis.
|
|
| 3. |
- Bludau, Isabell, et al.
(författare)
-
Systematic detection of functional proteoform groups from bottom-up proteomic datasets
- 2021
-
Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 12:1
-
Tidskriftsartikel (refereegranskat)abstract
- To a large extent functional diversity in cells is achieved by the expansion of molecular complexity beyond that of the coding genome. Various processes create multiple distinct but related proteins per coding gene – so-called proteoforms – that expand the functional capacity of a cell. Evaluating proteoforms from classical bottom-up proteomics datasets, where peptides instead of intact proteoforms are measured, has remained difficult. Here we present COPF, a tool for COrrelation-based functional ProteoForm assessment in bottom-up proteomics data. It leverages the concept of peptide correlation analysis to systematically assign peptides to co-varying proteoform groups. We show applications of COPF to protein complex co-fractionation data as well as to more typical protein abundance vs. sample data matrices, demonstrating the systematic detection of assembly- and tissue-specific proteoform groups, respectively, in either dataset. We envision that the presented approach lays the foundation for a systematic assessment of proteoforms and their functional implications directly from bottom-up proteomic datasets.
|
|
| 4. |
- Heusel, Moritz, et al.
(författare)
-
A Global Screen for Assembly State Changes of the Mitotic Proteome by SEC-SWATH-MS
- 2020
-
Ingår i: Cell systems. - : Elsevier BV. - 2405-4712. ; 10:2, s. 6-155
-
Tidskriftsartikel (refereegranskat)abstract
- Living systems integrate biochemical reactions that determine the functional state of each cell. Reactions are primarily mediated by proteins. In proteomic studies, these have been treated as independent entities, disregarding their higher-level organization into complexes that affects their activity and/or function and is thus of great interest for biological research. Here, we describe the implementation of an integrated technique to quantify cell-state-specific changes in the physical arrangement of protein complexes concurrently for thousands of proteins and hundreds of complexes. Applying this technique to a comparison of human cells in interphase and mitosis, we provide a systematic overview of mitotic proteome reorganization. The results recall key hallmarks of mitotic complex remodeling and suggest a model of nuclear pore complex disassembly, which we validate by orthogonal methods. To support the interpretation of quantitative SEC-SWATH-MS datasets, we extend the software CCprofiler and provide an interactive exploration tool, SECexplorer-cc.
|
|
| 5. |
- Hua, Kuo-Tai, et al.
(författare)
-
N-α-acetyltransferase 10 protein suppresses cancer cell metastasis by binding PIX proteins and inhibiting Cdc42/Rac1 activity
- 2011
-
Ingår i: Cancer Cell. - : Cell Press. - 1535-6108 .- 1878-3686. ; 19:2, s. 218-231
-
Tidskriftsartikel (refereegranskat)abstract
- N-α-acetyltransferase 10 protein, Naa10p, is an N-acetyltransferase known to be involved in cell cycle control. We found that Naa10p was expressed lower in varieties of malignancies with lymph node metastasis compared with non-lymph node metastasis. Higher Naa10p expression correlates the survival of lung cancer patients. Naa10p significantly suppressed migration, tumor growth, and metastasis independent of its enzymatic activity. Instead, Naa10p binds to the GIT-binding domain of PIX, thereby preventing the formation of the GIT-PIX-Paxillin complex, resulting in reduced intrinsic Cdc42/Rac1 activity and decreased cell migration. Forced expression of PIX in Naa10-transfected tumor cells restored the migration and metastasis ability. We suggest that Naa10p functions as a tumor metastasis suppressor by disrupting the migratory complex, PIX-GIT- Paxillin, in cancer cells.
|
|
| 6. |
- Malmstrom, Lars, et al.
(författare)
-
Automated Workflow for Large-Scale Selected Reaction Monitoring Experiments
- 2012
-
Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 11:3, s. 1644-1653
-
Tidskriftsartikel (refereegranskat)abstract
- Targeted proteomics allows researchers to study proteins of interest without being drowned in data from other, less interesting proteins or from redundant or uninformative peptides. While the technique is mostly used for smaller, focused studies, there are several reasons to conduct larger targeted experiments. Automated, highly robust software becomes more important in such experiments. In addition, larger experiments are carried out over longer periods of time, requiring strategies to handle the sometimes large shift in retention time often observed. We present a complete proof-of-principle software stack that automates most aspects of selected reaction monitoring workflows, a targeted proteomics technology. The soft-ware allows experiments to be easily designed and carried out. The steps automated are the generation of assays, generation of mass spectrometry driver files and methods files, and the import. and analysis of the data. All data are normalized to a common retention time scale, the data are then scored using a novel score model, and the error is subsequently estimated. We also show that selected reaction monitoring can be used for label-free quantification. All data generated are stored in a relational database, and the growing resource further facilitates the design of new experiments. We apply the technology to a large-scale experiment studying how Streptococcus pyogenes remodels its proteome under stimulation of human plasma.
|
|
| 7. |
|
|
| 8. |
- Teleman, Johan, et al.
(författare)
-
DIANA - algorithmic improvements for analysis of data-independent acquisition MS data.
- 2015
-
Ingår i: Bioinformatics. - : Oxford University Press (OUP). - 1367-4803 .- 1367-4811. ; 31:4, s. 555-562
-
Tidskriftsartikel (refereegranskat)abstract
- Data independent acquisition mass spectrometry has emerged as a reproducible and sensitive alternative in quantitative proteomics, where parsing the highly complex tandem mass spectra requires dedicated algorithms. Recently, targeted data extraction was proposed as a novel analysis strategy for this type of data, but it is important to further develop these concepts to provide quality-controlled, interference-adjusted and sensitive peptide quantification.
|
|
| 9. |
- Wisniewska, Magdalena, et al.
(författare)
-
Functional and Structural Properties of a Novel Protein and Virulence Factor (sHIP) in Streptococcus pyogenes.
- 2014
-
Ingår i: Journal of Biological Chemistry. - 1083-351X. ; 289:26, s. 18175-18188
-
Tidskriftsartikel (refereegranskat)abstract
- Streptococcus pyogenes is a significant bacterial pathogen in the human population. The importance of virulence factors for the survival and colonization of S. pyogenes is well established, and many of these factors are exposed to the extracellular environment enabling bacterial interactions with the host. In the present study we quantitatively analyzed and compared S. pyogenes proteins in the growth medium of a strain that is virulent to mice, with a non-virulent strain. Particularly one of these proteins was present at significantly higher levels in stationary growth medium from the virulent strain. We determined the three-dimensional structure of the protein that showed a unique tetrameric organization composed of four helix-loop-helix motifs. Affinity pull-down mass spectrometry analysis in human plasma demonstrated that the protein interacts with histidine-rich glycoprotein (HRG), and the name sHIP (streptococcal Histidine-rich glycoprotein Interacting Protein) is therefore proposed. HRG has antibacterial activity, and when challenged by HRG, sHIP was found to rescue S. pyogenes bacteria. This and the finding that patients with invasive S. pyogenes infection respond with antibody production against sHIP, suggest a role for the protein in S. pyogenes pathogenesis.
|
|
