Molecular & Cellular Proteomics最新文献

{"title":"Super Enhanced Purification of Denatured-Refolded Ubiquitinated Proteins by ThUBD Revealed Ubiquitinome Dysfunction in Liver Fibrosis.","authors":"Xinyu Cheng, Yonghong Wang, Jinfang Liu, Ying Wu, Zhenpeng Zhang, Hui Liu, Lantian Tian, Li Zhang, Lei Chang, Ping Xu, Lingqiang Zhang, Yanchang Li","doi":"10.1016/j.mcpro.2024.100852","DOIUrl":"10.1016/j.mcpro.2024.100852","url":null,"abstract":"<p><p>Ubiquitination is crucial for maintaining protein homeostasis and plays a vital role in diverse biological processes. Ubiquitinome profiling and quantification are of great scientific significance. Artificial ubiquitin-binding domains (UBDs) have been widely employed to capture ubiquitinated proteins. The success of this enrichment relies on recognizing native spatial structures of ubiquitin and ubiquitin chains by UBDs under native conditions. However, the use of native lysis conditions presents significant challenges, including insufficient protein extraction, heightened activity of deubiquitinating enzymes and proteasomes in removing the ubiquitin signal, and purification of a substantial number of contaminant proteins, all of which undermine the robustness and reproducibility of ubiquitinomics. In this study, we introduced a novel approach that combines denatured-refolded ubiquitinated sample preparation (DRUSP) with a tandem hybrid UBD for ubiquitinomic analysis. The samples were effectively extracted using strongly denatured buffers and subsequently refolded using filters. DRUSP yielded a significantly stronger ubiquitin signal, nearly three times greater than that of the Control method. Then, eight types of ubiquitin chains were quickly and accurately restored; therefore, they were recognized and enriched by tandem hybrid UBD with high efficiency and no biases. Compared with the Control method, DRUSP showed extremely high efficiency in enriching ubiquitinated proteins, improving overall ubiquitin signal enrichment by approximately 10-fold. Moreover, when combined with ubiquitin chain-specific UBDs, DRUSP had also been proven to be a versatile approach. This new method significantly enhanced the stability and reproducibility of ubiquitinomics research. Finally, DRUSP was successfully applied to deep ubiquitinome profiling of early mouse liver fibrosis with increased accuracy, revealing novel insights for liver fibrosis research.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100852"},"PeriodicalIF":6.1,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11584597/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142372343","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multilevel Proteomics Reveals Epigenetic Signatures in BCG-Mediated Macrophage Activation.","authors":"Zoe Schaefer, John Iradukunda, Evelyn N Lumngwena, Kari B Basso, Jonathan M Blackburn, Ivana K Parker","doi":"10.1016/j.mcpro.2024.100851","DOIUrl":"10.1016/j.mcpro.2024.100851","url":null,"abstract":"<p><p>The bacillus Calmette-Guérin BCG vaccine (Mycobacterium bovis) is primarily used to prevent tuberculosis (TB) infections but has wide-ranging immunogenic effects. One of its most notable properties is its ability to induce trained immunity, a memory-like response in innate immune cells such as macrophages. Through targeted analyses of well-established histone marks, prior research has shown that these changes are generated through epigenetic modification. Mass spectrometry-based proteomic approaches provide a way to globally profile various aspects of the proteome, providing data to further identify unexplored mechanisms of BCG-mediated immunomodulation. Here we use multi-level proteomics (total, histone, and phospho to identify networks and potential mechanisms that mediate BCG-induced immunomodulation in macrophages. Histone-focused proteomics and total proteomics were performed at the University of Cape Town (data available via ProteomeXchange with identifier PXD051187), while phosphoproteomics data was retrieved from the ProteomeXchange Repository (identifier PXD013171). We identify several epigenetic mechanisms that may drive BCG-induced training phenotypes. Evidence across the proteomics and histone-focused proteomics data set pair 6 epigenetic effectors (NuA4, NuRD, NSL, Sin3A, SIRT2, SIRT6) and their substrates.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100851"},"PeriodicalIF":6.1,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11585779/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142375632","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Proteome Dynamics in iPSC-Derived Human Dopaminergic Neurons.","authors":"Claudia Cavarischia-Rega, Karan Sharma, Julia C Fitzgerald, Boris Macek","doi":"10.1016/j.mcpro.2024.100838","DOIUrl":"10.1016/j.mcpro.2024.100838","url":null,"abstract":"<p><p>Dopaminergic neurons participate in fundamental physiological processes and are the cell type primarily affected in Parkinson's disease. Their analysis is challenging due to the intricate nature of their function, involvement in diverse neurological processes, and heterogeneity and localization in deep brain regions. Consequently, most of the research on the protein dynamics of dopaminergic neurons has been performed in animal cells ex vivo. Here we use iPSC-derived human mid-brain-specific dopaminergic neurons to study general features of their proteome biology and provide datasets for protein turnover and dynamics, including a human axonal translatome. We cover the proteome to a depth of 9409 proteins and use dynamic SILAC to measure the half-life of more than 4300 proteins. We report uniform turnover rates of conserved cytosolic protein complexes such as the proteasome and map the variable rates of turnover of the respiratory chain complexes in these cells. We use differential dynamic SILAC labeling in combination with microfluidic devices to analyze local protein synthesis and transport between axons and soma. We report 105 potentially novel axonal markers and detect translocation of 269 proteins between axons and the soma in the time frame of our analysis (120 h). Importantly, we provide evidence for local synthesis of 154 proteins in the axon and their retrograde transport to the soma, among them several proteins involved in RNA editing such as ADAR1 and the RNA helicase DHX30, involved in the assembly of mitochondrial ribosomes. Our study provides a workflow and resource for the future applications of quantitative proteomics in iPSC-derived human neurons.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100838"},"PeriodicalIF":6.1,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11474371/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142291445","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identification of a Proteomic Signature for Predicting Immunotherapy Response in Patients With Metastatic Non-Small Cell Lung Cancer.","authors":"Patricia Mondelo-Macía, Jorge García-González, Luis León-Mateos, Alicia Abalo, Susana Bravo, María Del Pilar Chantada Vazquez, Laura Muinelo-Romay, Rafael López-López, Roberto Díaz-Peña, Ana B Dávila-Ibáñez","doi":"10.1016/j.mcpro.2024.100834","DOIUrl":"10.1016/j.mcpro.2024.100834","url":null,"abstract":"<p><p>Immunotherapy has improved survival rates in patients with cancer, but identifying those who will respond to treatment remains a challenge. Advances in proteomic technologies have enabled the identification and quantification of nearly all expressed proteins in a single experiment. Integrating mass spectrometry with high-throughput technologies has facilitated comprehensive analysis of the plasma proteome in cancer, facilitating early diagnosis and personalized treatment. In this context, our study aimed to investigate the predictive and prognostic value of plasma proteome analysis using the SWATH-MS (Sequential Window Acquisition of All Theoretical Mass Spectra) strategy in newly diagnosed patients with non-small cell lung cancer (NSCLC) receiving pembrolizumab therapy. We enrolled 64 newly diagnosed patients with advanced NSCLC treated with pembrolizumab. Blood samples were collected from all patients before and during therapy. A total of 171 blood samples were analyzed using the SWATH-MS strategy. Plasma protein expression in metastatic NSCLC patients prior to receiving pembrolizumab was analyzed. A first cohort (discovery cohort) was employed to identify a proteomic signature predicting immunotherapy response. Thus, 324 differentially expressed proteins between responder and non-responder patients were identified. In addition, we developed a predictive model and found a combination of seven proteins, including ATG9A, DCDC2, HPS5, FIL1L, LZTL1, PGTA, and SPTN2, with stronger predictive value than PD-L1 expression alone. Additionally, survival analyses showed an association between the levels of ATG9A, DCDC2, SPTN2 and HPS5 with progression-free survival (PFS) and/or overall survival (OS). Our findings highlight the potential of proteomic technologies to detect predictive biomarkers in blood samples from NSCLC patients, emphasizing the correlation between immunotherapy response and the idenfied protein set.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100834"},"PeriodicalIF":6.1,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11474190/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142109572","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Nitrogen-specific Interactome Analysis Sheds Light on the Role of the SnRK1 and TOR Kinases in Plant Nitrogen Signaling.","authors":"Freya Persyn, Wouter Smagghe, Dominique Eeckhout, Toon Mertens, Thomas Smorscek, Nancy De Winne, Geert Persiau, Eveline Van De Slijke, Nathalie Crepin, Astrid Gadeyne, Jelle Van Leene, Geert De Jaeger","doi":"10.1016/j.mcpro.2024.100842","DOIUrl":"10.1016/j.mcpro.2024.100842","url":null,"abstract":"<p><p>Nitrogen (N) is of utmost importance for plant growth and development. Multiple studies have shown that N signaling is tightly coupled with carbon (C) levels, but the interplay between C/N metabolism and growth remains largely an enigma. Nonetheless, the protein kinases Sucrose Non-fermenting 1 (SNF1)-Related Kinase 1 (SnRK1) and Target Of Rapamycin (TOR), two ancient central metabolic regulators, are emerging as key integrators that link C/N status with growth. Despite their pivotal importance, the exact mechanisms behind the sensing of N status and its integration with C availability to drive metabolic decisions are largely unknown. Especially for SnRK1, it is not clear how this kinase responds to altered N levels. Therefore, we first monitored N-dependent SnRK1 kinase activity with an in vivo Separation of Phase-based Activity Reporter of Kinase (SPARK) sensor, revealing a contrasting N-dependency in Arabidopsis thaliana (Arabidopsis) shoot and root tissues. Next, using affinity purification (AP) and proximity labeling (PL) coupled to mass spectrometry (MS) experiments, we constructed a comprehensive SnRK1 and TOR interactome in Arabidopsis cell cultures during N-starved and N-repleted growth conditions. To broaden our understanding of the N-specificity of the TOR/SnRK1 signaling events, the resulting network was compared to corresponding C-related networks, identifying a large number of novel, N-specific interactors. Moreover, through integration of N-dependent transcriptome and phosphoproteome data, we were able to pinpoint additional N-dependent network components, highlighting for instance SnRK1 regulatory proteins that might function at the crosstalk of C/N signaling. Finally, confirmation of known and identification of novel SnRK1 interactors, such as Inositol-Requiring 1 (IRE1A) and the RAB GTPase RAB18, indicate that SnRK1, present at the ER, is involved in N signaling and autophagy induction.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100842"},"PeriodicalIF":6.1,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11526089/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142291441","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Combining Data Independent Acquisition With Spike-In SILAC (DIA-SiS) Improves Proteome Coverage and Quantification.","authors":"Anna Sophie Welter, Maximilian Gerwien, Robert Kerridge, Keziban Merve Alp, Philipp Mertins, Matthias Selbach","doi":"10.1016/j.mcpro.2024.100839","DOIUrl":"10.1016/j.mcpro.2024.100839","url":null,"abstract":"<p><p>Data-independent acquisition (DIA) is increasingly preferred over data-dependent acquisition due to its higher throughput and fewer missing values. Whereas data-dependent acquisition often uses stable isotope labeling to improve quantification, DIA mostly relies on label-free approaches. Efforts to integrate DIA with isotope labeling include chemical methods like mass differential tags for relative and absolute quantification and dimethyl labeling, which, while effective, complicate sample preparation. Stable isotope labeling by amino acids in cell culture (SILAC) achieves high labeling efficiency through the metabolic incorporation of heavy labels into proteins in vivo. However, the need for metabolic incorporation limits the direct use in clinical scenarios and certain high-throughput experiments. Spike-in SILAC (SiS) methods use an externally generated heavy sample as an internal reference, enabling SILAC-based quantification even for samples that cannot be directly labeled. Here, we combine DIA-SiS, leveraging the robust quantification of SILAC without the complexities associated with chemical labeling. We developed DIA-SiS and rigorously assessed its performance with mixed-species benchmark samples on bulk and single cell-like amount level. We demonstrate that DIA-SiS substantially improves proteome coverage and quantification compared to label-free approaches and reduces incorrectly quantified proteins. Additionally, DIA-SiS proves effective in analyzing proteins in low-input formalin-fixed paraffin-embedded tissue sections. DIA-SiS combines the precision of stable isotope-based quantification with the simplicity of label-free sample preparation, facilitating simple, accurate, and comprehensive proteome profiling.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100839"},"PeriodicalIF":6.1,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11795695/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142291442","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Proteomic Heterogeneity of the Extracellular Matrix Identifies Histologic Subtype-Specific Fibroblast in Gastric Cancer.","authors":"Hyun Jin Lee, Yoonjin Kwak, Yun Suk Na, Hyejin Kim, Mi Ree Park, Jeong Yeon Jo, Jin Young Kim, Soo-Jeong Cho, Pilnam Kim","doi":"10.1016/j.mcpro.2024.100843","DOIUrl":"10.1016/j.mcpro.2024.100843","url":null,"abstract":"<p><p>Gastric cancer (GC) is a highly heterogeneous disease regarding histologic features, genotypes, and molecular phenotypes. Here, we investigate extracellular matrix (ECM)-centric analysis, examining its association with histologic subtypes and patient prognosis in human GC. We performed quantitative proteomic analysis of decellularized GC tissues that characterizes tumorous ECM, highlighting proteomic heterogeneity in ECM components. We identified 20 tumor-enriched proteins including four glycoproteins, serpin family H member 1 (SERPINH1), annexin family (ANXA3/4/5/13), S100A family (S100A6/8/9), MMP14, and other matrisome-associated proteins. In addition, histopathological characteristics of GC reveals differential expression in ECM composition, with the poorly cohesive carcinoma-not otherwise specified (PCC-NOS) subtype being distinctly demarcated from other histologic subtypes. Integrating ECM proteomics with single-cell RNA sequencing, we identified crucial molecular markers in the PCC-NOS-specific stroma. PCC-NOS-enriched matrisome proteins and gene expression signatures of adipogenic cancer-associated fibroblasts (CAF_adi) are closely linked, both associated with adverse outcomes in GC. Using tumor microarray analysis, we confirmed the CAF_adi surface marker, ATP binding cassette subfamily A member 8 (ABCA8), predominantly present in PCC-NOS tumors. Our ECM-focused analysis paves the way for studies to determine their utility as biomarkers for patient stratification, offering valuable insights for linking molecular and histologic features in GC.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100843"},"PeriodicalIF":6.1,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11526087/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142291447","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Universal Identification of Pathogenic Viruses by Liquid Chromatography Coupled with Tandem Mass Spectrometry Proteotyping.","authors":"Clément Lozano, Olivier Pible, Marine Eschlimann, Mathieu Giraud, Stéphanie Debroas, Jean-Charles Gaillard, Laurent Bellanger, Laurent Taysse, Jean Armengaud","doi":"10.1016/j.mcpro.2024.100822","DOIUrl":"10.1016/j.mcpro.2024.100822","url":null,"abstract":"<p><p>Accurate and rapid identification of viruses is crucial for an effective medical diagnosis when dealing with infections. Conventional methods, including DNA amplification techniques or lateral-flow assays, are constrained to a specific set of targets to search for. In this study, we introduce a novel tandem mass spectrometry proteotyping-based method that offers a universal approach for the identification of pathogenic viruses and other components, eliminating the need for a priori knowledge of the sample composition. Our protocol relies on a time and cost-efficient peptide sample preparation, followed by an analysis with liquid chromatography coupled to high-resolution tandem mass spectrometry. As a proof of concept, we first assessed our method on publicly available shotgun proteomics datasets obtained from virus preparations and fecal samples of infected individuals. Successful virus identification was achieved with 53 public datasets, spanning 23 distinct viral species. Furthermore, we illustrated the method's capability to discriminate closely related viruses within the same sample, using alphaviruses as an example. The clinical applicability of our method was demonstrated by the accurate detection of the vaccinia virus in spiked saliva, a matrix of paramount clinical significance due to its non-invasive and easily obtainable nature. This innovative approach represents a significant advancement in pathogen detection and paves the way for enhanced diagnostic capabilities.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100822"},"PeriodicalIF":6.1,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11795680/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141860279","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"ZASP: A Highly Compatible and Sensitive ZnCl₂ Precipitation-Assisted Sample Preparation Method for Proteomic Analysis.","authors":"Xianfeng Shao, Yuanxuan Huang, Rong Xu, Qiqing He, Min Zhang, Fuchu He, Dongxue Wang","doi":"10.1016/j.mcpro.2024.100837","DOIUrl":"10.1016/j.mcpro.2024.100837","url":null,"abstract":"<p><p>Universal sample preparation for proteomic analysis that enables unbiased protein manipulation, flexible reagent use, and low protein loss is required to ensure the highest sensitivity of downstream liquid chromatography-mass spectrometry (LC-MS) analysis. To address these needs, we developed a ZnCl₂ precipitation-assisted sample preparation method (ZASP) that depletes harsh detergents and impurities in protein solutions prior to trypsin digestion via 10 min of ZnCl₂ and methanol-induced protein precipitation at room temperature (RT). ZASP can remove trypsin digestion and LC-MS incompatible detergents such as SDS, Triton X-100, and urea at high concentrations in solution and unbiasedly recover proteins independent of the amount of protein input. We demonstrated the sensitivity and reproducibility of ZASP in an analysis of samples with 1 μg to 1000 μg of proteins. Compared to commonly used sample preparation methods such as SDC-based in-solution digestion, acetone precipitation, FASP, and SP3, ZASP has proven to be an efficient approach. Here, we present ZASP, a practical, robust, and cost-effective proteomic sample preparation method that can be applied to profile different types of samples.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100837"},"PeriodicalIF":6.1,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11492125/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142145981","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"metaExpertPro: A Computational Workflow for Metaproteomics Spectral Library Construction and Data-Independent Acquisition Mass Spectrometry Data Analysis.","authors":"Yingying Sun, Ziyuan Xing, Shuang Liang, Zelei Miao, Lai-Bao Zhuo, Wenhao Jiang, Hui Zhao, Huanhuan Gao, Yuting Xie, Yan Zhou, Liang Yue, Xue Cai, Yu-Ming Chen, Ju-Sheng Zheng, Tiannan Guo","doi":"10.1016/j.mcpro.2024.100840","DOIUrl":"10.1016/j.mcpro.2024.100840","url":null,"abstract":"<p><p>Analysis of large-scale data-independent acquisition mass spectrometry metaproteomics data remains a computational challenge. Here, we present a computational pipeline called metaExpertPro for metaproteomics data analysis. This pipeline encompasses spectral library generation using data-dependent acquisition MS, protein identification and quantification using data-independent acquisition mass spectrometry, functional and taxonomic annotation, as well as quantitative matrix generation for both microbiota and hosts. By integrating FragPipe and DIA-NN, metaExpertPro offers compatibility with both Orbitrap and timsTOF MS instruments. To evaluate the depth and accuracy of identification and quantification, we conducted extensive assessments using human fecal samples and benchmark tests. Performance tests conducted on human fecal samples indicated that metaExpertPro quantified an average of 45,000 peptides in a 60-min diaPASEF injection. Notably, metaExpertPro outperformed three existing software tools by characterizing a higher number of peptides and proteins. Importantly, metaExpertPro maintained a low factual false discovery rate of approximately 5% for protein groups across four benchmark tests. Applying a filter of five peptides per genus, metaExpertPro achieved relatively high accuracy (F-score = 0.67-0.90) in genus diversity and showed a high correlation (r_Spearman = 0.73-0.82) between the measured and true genus relative abundance in benchmark tests. Additionally, the quantitative results at the protein, taxonomy, and function levels exhibited high reproducibility and consistency across the commonly adopted public human gut microbial protein databases IGC and UHGP. In a metaproteomic analysis of dyslipidemia patients, metaExpertPro revealed characteristic alterations in microbial functions and potential interactions between the microbiota and the host.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100840"},"PeriodicalIF":6.1,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11795700/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142291444","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}