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  • Liquid Biopsy‐Based Single‐Cell Transcriptome Profiling Characterizes Heterogeneity of Disseminated Tumor Cells from Lung Adenocarcinoma
    Proteomics (IF 3.106) Pub Date : 2020-01-21
    Yu Dong; Zhuo Wang; Qihui Shi

    The advent of rapid and inexpensive sequencing technology allows scientists to decipher intratumor heterogeneity spatially and temporally for resolving the evolutionary history of tumor and the underlying mechanism. However, studies on characterizing heterogeneity of disseminated tumor cells (DTCs) in liquid biopsies are rare because of the rarity and low viability of DTCs as well as a large number non‐tumor cells. Here, we employed high‐throughput single‐cell transcriptome sequencing technology and rare DTC enrichment method to decipher the heterogeneity and distinct molecular signatures of DTCs in malignant pleural effusion (MPE) from lung adenocarcinoma. Single‐cell transcriptomes of 8,213 MPE‐derived cells were acquired for bioinformatics analysis. In these cells from MPE, five main cell populations including tumor, mesothelial, monocyte, T and B cells were identified with specific markers for each group. Tumor cells present in MPE were further divided into four distinct subgroups that were found to be associated with immune response, cell proliferation, apoptosis and cell adhesion, respectively. Based on the single‐cell dataset of MPE‐derived DTCs, we identified 19 tumor‐specific markers that were also highly expressed at RNA and protein levels in tumor tissues as candidate markers for detection of extraordinarily rare circulating tumor cells in the blood.

    更新日期:2020-01-22
  • Emerging Roles and Research Tools of Atypical Ubiquitination
    Proteomics (IF 3.106) Pub Date : 2020-01-12
    Qiuling Huang; Xiaofei Zhang

    Ubiquitination is a posttranslational modification (PTM) characterized by the covalent attachment of ubiquitin molecules to protein substrates. The ubiquitination modification process is reversible, dynamic and involved in the regulation of various biological processes, such as autophagy, inflammatory responses, and DNA damage responses. The forms of ubiquitin modification are very diverse, incorporating either a single ubiquitin molecule or a complicated ubiquitin polymer, and different types of ubiquitination usually elicit corresponding cellular responses. The development of research tools and strategies has afforded more detailed insight into atypical ubiquitin signaling pathways that were previously poorly understood. Here, we provide an update on the understanding of atypical ubiquitin chain signaling pathways and discuss the recent development of representative research tools for ubiquitin systems. In addition, we reflect on and summarize the future challenges in ubiquitin research.

    更新日期:2020-01-13
  • Profiling the Protein Targets of Unmodified Bio‐Active Molecules with Drug Affinity Responsive Target Stability and Liquid Chromatography/Tandem Mass Spectrometry
    Proteomics (IF 3.106) Pub Date : 2020-01-11
    Hui‐Yun Hwang; Tae Young Kim; Marcell A. Szász; Balazs Dome; Johan Malm; Gyorgy Marko‐Varga; Ho Jeong Kwon

    Identifying the target proteins of bioactive small molecules is a key step in understanding mode‐of‐action of the drug and addressing the underlying mechanisms responsible for a particular phenotype. Proteomics has been successfully used to elucidate the target protein profiles of unmodified and ligand‐modified bioactive small molecules. In the latter approach, compounds can be modified via click chemistry and combined with activity‐based protein profiling. Target proteins are then enriched by performing a pull‐down with the modified ligand. Methods that utilize unmodified bioactive small molecules include the cellular thermal shift assay, thermal proteome profiling, stability of proteins from rates of oxidation, and the drug affinity responsive target stability determination (or read‐out). This review highlights recent proteomic approaches utilizing data‐dependent analysis and data independent analysis to identify target proteins by DARTS. When combined with LC‐MS/MS, DARTS enables the identification of proteins that bind to drug molecules that leads to a conformational change in the target protein(s). In addition, we propose an effective strategy for selecting the target protein(s) from within the pool of analyzed candidates. With additional complementary methods, the biologically‐relevant target proteins that bind to the small bio‐active molecules can be further validated.

    更新日期:2020-01-13
  • Streamlined Development of Targeted Mass Spectrometry‐Based Method Combining Scout‐MRM and a Web‐Based Tool Indexed with Scout Peptides
    Proteomics (IF 3.106) Pub Date : 2020-01-10
    Sophie Ayciriex; Romain Carrière; Chloé Bardet; J. C. Yves Le Blanc; Arnaud Salvador; Tanguy Fortin; Jérôme Lemoine

    MS‐based targeted proteomics is a relevant technology for sensitive and robust relative or absolute quantification of proteins biomarker candidates in complex human biofluids or tissue extracts. Performing a multiplex assay imposes time scheduling of peptide monitoring only around their expected retention time that needs to be defined with synthetic peptide. Time‐scheduled monitoring is clearly a constraint that precludes from straightforward assay transfer between biological matrices or distinct experimental setup. Any unexpected retention time (RT) shift challenges assay robustness and its implementation for large‐scale analysis. Recently, Scout‐multiple reaction monitoring that fully releases multiplexed targeted acquisition from RT scheduling by successively monitoring complex transition groups triggered with sentinel molecules called Scout has been introduced. It is herein documented how Peptide Selector database and tool streamlines the building of a multiplexed method thanks to RT indexation relative to Scout peptides. This case study deals with surrogate peptides of biomarker candidates related to drug‐induced liver and vascular injury, running such on‐line built method (eight Scouts triggering the monitoring of a total of 692 transitions) enables 100% recovery of a panel of 93 spiked‐in heavy labeled standards, despite significant RT shifts between serum, plasma, or urine. This result illustrates the simplicity of automatically building and deploying robust proteomics targeted assay.

    更新日期:2020-01-11
  • Data‐Independent Acquisition‐Based Quantitative Proteomics Analysis Reveals Dynamic Network Profiles during the Macrophage Inflammatory Response
    Proteomics (IF 3.106) Pub Date : 2020-01-10
    Lei Li; Li Chen; Xinya Lu; Chenyang Huang; Haihua Luo; Jingmiao Jin; Zhuzhong Mei; Jinghua Liu; Cuiting Liu; Junmin Shi; Peng Chen; Yong Jiang

    Understanding of the molecular regulatory mechanisms underlying the inflammatory response is incomplete. The present study focuses on characterizing the proteome in a model of inflammation in macrophages treated with lipopolysaccharide (LPS). A total of 3597 proteins are identified in macrophages with the data‐independent acquisition (DIA) method. Bioinformatic analyses reveal discrete modules and the underlying molecular mechanisms, as well as the signaling network that modulates the development of inflammation. It is found that a total of 87 differentially expressed proteins are shared by all stages of LPS‐induced inflammation in macrophages and that 18 of these proteins participate in metabolic processes by forming a tight interaction network. Data support the hypothesis that ribosome proteins play a key role in regulating the macrophage response to LPS. Interestingly, conjoint analyses of the transcriptome and proteome in macrophages treated with LPS reveal that the genes upregulated at both the mRNA and protein levels are mainly involved in inflammation and the immune response, whereas the genes downregulated are significantly enriched in metabolism‐related processes. These results not only provide a more comprehensive understanding of the molecular mechanisms of inflammation mediated by bacterial infection but also provide a dynamic proteomic resource for further studies.

    更新日期:2020-01-11
  • Mass Spectrometry Reveals New Insights into the Production of Superoxide Anions and 4‐Hydroxynonenal Adducted Proteins in Human Sperm
    Proteomics (IF 3.106) Pub Date : 2020-01-09
    Jacob Kendal Netherton; Louise Hetherington; Rachel Anne Ogle; Marum Mazloumi Gavgani; Tony Velkov; Ana Izabel Bilbin Villaverde; Nuch Tanphaichitr; Mark Andrew Baker

    The free‐radical theory of male infertility suggests that reactive oxygen species produced by the spermatozoa themselves are a leading cause of sperm dysfunction, including loss of sperm motility. However, the field is overshadowed on several fronts, primarily because: i) the probes used to measure reactive oxygen species (ROS) are imprecise; and ii) many reports suggesting that oxygen radicals are detrimental to sperm function add an exogenous source of ROS. Herein, a more reliable approach to measure superoxide anion production by human spermatozoa based on MS analysis is used. Furthermore, the formation of the lipid‐peroxidation product 4‐hydroxynonenal (4‐HNE) during in vitro incubation using proteomics is also investigated. The data demonstrate that neither superoxide anion nor other free radicals that cause 4‐HNE production are related to the loss of sperm motility during incubation. Interestingly, it appears that many of the 4‐HNE adducted proteins, found within spermatozoa, originate from the prostate. A quantitative SWATH analysis demonstrate that these proteins transiently bind to sperm and are then shed during in vitro incubation. These proteomics‐based findings propose a revised understanding of oxidative stress within the male reproductive tract.

    更新日期:2020-01-09
  • PDMS Microwell Stencil Based Multiplexed Single‐Cell Secretion Analysis
    Proteomics (IF 3.106) Pub Date : 2020-01-09
    Meimei Liu; Meihua Jin; Linmei Li; Yahui Ji; Fengjiao Zhu; Yong Luo; Tingjiao Liu; Bingcheng Lin; Yao Lu

    Multiplexed single‐cell protein secretion analysis provides an in‐depth understanding of cellular heterogeneity in intercellular communications mediated by secreted proteins in both fundamental and clinical research. However, it has been challenging to increase the proteomic parameters co‐profiled from every single cell in a facile way. Herein, a simple method to improve the multiplexed proteomic parameters of PDMS microwell based single‐cell secretion analysis platform by sandwiching PDMS stencil in between two antibody‐coated glass slides is introduced. Two different antibody panels can be immobilized easily by static coating, without using sophisticated fluid handling or bulky equipment. 5‐plexed, 3‐fluorescence color single‐cell secretion assay is demonstrated with this platform to investigate human monocytic U937 cells in response to lipopolysaccharide and phorbol myristate acetate stimulation, which identified the existence of functional subsets dictated by different cytokine profiles. The technology introduced here is simple, easy to operate, which holds great potential to become a powerful tool for profiling multiplexed single‐cell cytokine secretion at high throughput to dissect cellular heterogeneity in secretome signatures.

    更新日期:2020-01-09
  • Proteomics of Long‐Lived Mammals
    Proteomics (IF 3.106) Pub Date : 2020-01-09
    Gregory Tombline; Jonathan Gigas; Nicholas Macoretta; Max Zacher; Stephan Emmrich; Yang Zhao; Andrei Seluanov; Vera Gorbunova

    Mammalian species differ up to 100‐fold in their aging rates and maximum lifespans. Long‐lived mammals appear to possess traits that extend lifespan and healthspan. Genomic analyses have not revealed a single pro‐longevity function that would account for all longevity effects. In contrast, it appears that pro‐longevity mechanisms may be complex traits afforded by connections between metabolism and protein functions that are impossible to predict by genomic approaches alone. Thus, metabolomics and proteomics studies will be required to understand the mechanisms of longevity. Several examples are reviewed that demonstrate the naked mole rat (NMR) shows unique proteomic signatures that contribute to longevity by overcoming several hallmarks of aging. SIRT6 is also discussed as an example of a protein that evolves enhanced enzymatic function in long‐lived species. Finally, it is shown that several longevity‐related proteins such as Cip1/p21, FOXO3, TOP2A, AKT1, RICTOR, INSR, and SIRT6 harbor posttranslational modification (PTM) sites that preferentially appear in either short‐ or long‐lived species and provide examples of crosstalk between PTM sites. Prospects of enhancing lifespan and healthspan of humans by altering metabolism and proteoforms with drugs that mimic changes observed in long‐lived species are discussed.

    更新日期:2020-01-09
  • Dynamic Phosphoproteome Profiling of Zebrafish Embryonic Fibroblasts during Cold Acclimation
    Proteomics (IF 3.106) Pub Date : 2020-01-08
    Junjun Yan; Yong Long; Tong Zhou; Jing Ren; Qing Li; Guili Song; Zongbin Cui

    Temperature affects almost all aspects of the fish life. To cope with low temperature, fish have evolved the ability of cold acclimation for survival. However, intracellular signaling events underlying cold acclimation in fish remain largely unknown. Here, the formation of cold acclimation in zebrafish embryonic fibroblasts (ZF4) is monitored and the phosphorylation events during the process are investigated through a large‐scale quantitative phosphoproteomic approach. In total, 11 474 phosphorylation sites are identified on 4066 proteins and quantified 5772 phosphosites on 2519 proteins. Serine, threonine, and tyrosine (Ser/Thr/Tyr) phosphorylation accounted for 85.5%, 13.3%, and 1.2% of total phosphosites, respectively. Among all phosphosites, 702 phosphosites on 510 proteins show differential regulation during cold acclimation of ZF4 cells. These phosphosites are divided into six clusters according to their dynamic changes during cold exposure. Kinase–substrate prediction reveals that mitogen‐activated protein kinase (MAPK) among the kinase groups is predominantly responsible for phosphorylation of these phosphosites. The differentially regulated phosphoproteins are functionally associated with various cellular processes such as regulation of actin cytoskeleton and MAPK signaling pathway. These data enrich the database of protein phosphorylation sites in zebrafish and provide key clues for the elucidation of intracellular signaling networks during cold acclimation of fish.

    更新日期:2020-01-08
  • Label‐Free Quantitative Proteomics Distinguishes General and Site‐Specific Host Responses to Pseudomonas aeruginosa Infection at the Ocular Surface
    Proteomics (IF 3.106) Pub Date : 2020-01-07
    Jason Yeung; Mihaela Gadjeva; Jennifer Geddes‐McAlister

    Mass spectrometry‐based proteomics enables the unbiased and sensitive profiling of cellular proteomes and extracellular environments. Recent technological and bioinformatic advances permit identifying dual biological systems in a single experiment, supporting investigation of infection from both the host and pathogen perspectives. At the ocular surface, Pseudomonas aeruginosa is commonly associated with biofilm formation and inflammation of the ocular tissues, causing damage to the eye. The interaction between P. aeruginosa and the immune system at the site of infection describes limitations in clearance of infection and enhanced pathogenesis. Here, the extracellular environment (eye wash) of murine ocular surfaces infected with a clinical isolate of P. aeruginosa is profiled and neutrophil marker proteins are detected, indicating neutrophil recruitment to the site of infection. The first potential diagnostic markers of P. aeruginosa‐associated keratitis are also identified. In addition, the deepest murine corneal proteome to date is defined and proteins, categories, and networks critical to the host response are detected. Moreover, the first identification of bacterial proteins attached to the ocular surface is reported. The findings are validated through in silico comparisons and enzymatic profiling. Overall, the work provides comprehensive profiling of the host–pathogen interface and uncovers differences between general and site‐specific host responses to infection.

    更新日期:2020-01-07
  • Microfluidic Cell Trap Arrays for Single Hematopoietic Stem/Progenitor Cell Behavior Analysis
    Proteomics (IF 3.106) Pub Date : 2020-01-03
    Xin Han; Yuan Ma; Kai Zhang; Pengchao Zhang; Ning Shao; Lidong Qin

    Hematopoietic stem/progenitor cell (HSPC) mobilization from the bone marrow to the bloodstream is a required step for blood cell renewal, and HSPC motility is a clinically relevant standard for peripheral blood stem cell transplantation. Individual HSPCs exhibit considerable heterogeneity in motility behaviors, which are subject to complex intrinsic and extrinsic regulatory mechanisms. Motility‐based cell sorting is then demanded to fulfill the study of such mechanism complexity. However, due to the HSPC heterogeneity and difficulty in monitoring cell motility, such a platform is still not available. With the recent development of microfluidics technology, motility‐based monitoring, sorting, collecting, and analysis of HSPC behaviors are highly possible and achievable if fluid channels and structures are correctly engineered. Here, a new design of microfluidic arrays for single‐cell trapping is presented, enabling high‐throughput analysis of individual HSPC motility and behavior. Using these arrays, it is observed that HSPC motility is positively correlated with CD34 asymmetric inheritance and cell differentiation. Transcriptomic analysis of HSPCs sorted according to motility reveals changes in expression of genes associated with the regulation of stem‐cell maintenance. Ultimately, this novel, physical cell‐sorting system can facilitate the screening of HSPC mobilization compounds and the analysis of signals driving HSPC fate decisions.

    更新日期:2020-01-04
  • Calcitriol Prevents RAD51 Loss and cGAS‐STING‐IFN Response Triggered by Progerin
    Proteomics (IF 3.106) Pub Date : 2019-12-30
    Nuria Coll‐Bonfill; Rafael Cancado de Faria; Sweta Bhoopatiraju; Susana Gonzalo

    Hutchinson Gilford progeria syndrome (HGPS) is a devastating accelerated aging disease caused by LMNA gene mutation. The truncated lamin A protein produced “progerin” has a dominant toxic effect in cells, causing disruption of nuclear architecture and chromatin structure, genomic instability, gene expression changes, oxidative stress, and premature senescence. It was previously shown that progerin‐induced genomic instability involves replication stress (RS), characterized by replication fork stalling and nuclease‐mediated degradation of stalled forks. RS is accompanied by activation of cGAS/STING cytosolic DNA sensing pathway and STAT1‐regulated interferon (IFN)‐like response. It is also found that calcitriol, the active hormonal form of vitamin D, rescues RS and represses the cGAS/STING/IFN cascade. Here, the mechanisms underlying RS in progerin‐expressing cells and the rescue by calcitriol are explored. It is found that progerin elicits a marked downregulation of RAD51, concomitant with increased levels of phosphorylated‐RPA, a marker of RS. Interestingly, calcitriol prevents RS and activation of the cGAS/STING/IFN response in part through maintenance of RAD51 levels in progerin‐expressing cells. Thus, loss of RAD51 is one of the consequences of progerin expression that can contribute to RS and activation of the IFN response. Stabilization of RAD51 helps explain the beneficial effects of calcitriol in these processes.

    更新日期:2019-12-30
  • Pressure Cycling Technology‐Assisted Mass Spectrometric Quantification of Gingival Tissue Reveals Proteome Dynamics During the Initiation and Progression of Inflammatory Periodontal Disease
    Proteomics (IF 3.106) Pub Date : 2019-12-27
    Kai Bao; Xiaofei Li; Tetsuhiro Kajikawa; Abe Toshiharu; Nathalie Selevsek; Jonas Grossmann; George Hajishengallis; Nagihan Bostanci

    Understanding the progression of periodontal tissue destruction is at the forefront of periodontal research. We aimed to capture the dynamics of gingival tissue proteome during the initiation and progression of experimental (ligature‐induced) periodontitis in mice. Pressure cycling technology (PCT), a recently developed platform that uses ultra‐high pressure to disrupt tissues, was utilized to achieve efficient and reproducible protein extraction from ultra‐small amounts of gingival tissues in combination with liquid chromatography‐tandem mass spectrometry (MS). The MS data were processed using Progenesis QI and the regulated proteins were subjected to METACORE, STRING, and WebGestalt for functional enrichment analysis. A total of 1614 proteins with ≥2 peptides were quantified with an estimated protein false discovery rate of 0.06%. Unsupervised clustering analysis showed that the gingival tissue protein abundance is mainly dependent on the periodontitis progression stage. Gene ontology enrichment analysis revealed an over‐representation in innate immune regulation (e.g., neutrophil‐mediated immunity and antimicrobial peptides), signal transduction (e.g., integrin signaling), and homeostasis processes (e.g., platelet activation and aggregation). In conclusion, we applied a PCT‐assisted label‐free quantitative proteomics workflow that allowed cataloging the deepest gingival tissue proteome on a rapid timescale and provided novel mechanistic insights into host perturbation during periodontitis progression.

    更新日期:2019-12-29
  • Comparative proteomic profiling of methicillin‐susceptible and resistant Staphylococcus aureus
    Proteomics (IF 3.106) Pub Date : 2019-12-24
    Zhen Xu; Jiazhen Chen; Kostas Vougas; Ajit Shah; Haroun Shah; Raju Misra; Hermine V Mkrtchyan

    Staphylococcus aureus is a highly successful human pathogen responsible for wide range of infections. In this study, we provide insights into the virulence, pathogenicity, and antimicrobial resistance determinants of methicillin susceptible and methicillin resistant Staphylococcus aureus (MSSA; MRSA) recovered from non‐healthcare environments.

    更新日期:2019-12-25
  • Proteomic Profile of Human Schwann Cells
    Proteomics (IF 3.106) Pub Date : 2019-12-23
    Aysha Ferdoushi; Xiang Li; Muhammad Fairuz Bin Jamaluddin; Hubert Hondermarck

    Schwann cells (SC) are essential for the growth, maintenance, and regeneration of peripheral nerves, but the proteome of normal human SC is poorly defined. Here, a proteomic analysis by LC–MS/MS is performed to define the protein expression profile of primary human SC. A total of 19 557 unique peptides corresponding to 1553 individual proteins are identified. Ingenuity Pathway Analysis (IPA), Gene Ontology (GO), and Database for Annotation, Visualization, and Integrated Discovery (DAVID) are used to assign protein localization and function, and to define enriched pathways. EIF2, mTOR, and integrin signaling are among the most enriched pathways and the most enriched biological function is cell–cell adhesion, which is in agreement with the supportive role of SC in peripheral nerves. In addition, several nociceptors and synaptic proteins are identified and may contribute to the recently discovered role of SC in pain sensation and cancer progression. This proteome analysis of normal human SC constitutes a reference for future molecular explorations of physiological and pathological processes where SC are involved.

    更新日期:2019-12-23
  • Neurogranin Regulates Alcohol Sensitivity through AKT Pathway in the Nucleus Accumbens
    Proteomics (IF 3.106) Pub Date : 2019-12-23
    Hyung W. Nam; Caleb A. Grant; Ashton N. Jorgensen; Carrie J. Holtz‐Heppelmann; Marjan Trutschl; Urska Cvek

    Dysfunction of glutamate neurotransmission in the nucleus accumbens (NAc) has been implicated in the pathophysiology of alcohol use disorders (AUD). Neurogranin (Ng) is exclusively expressed in the brain and mediates N‐methyl‐d‐aspartate receptor (NMDAR) hypo‐function by regulating the intracellular calcium‐calmodulin (Ca2+‐CaM) pathway. Ng null mice (Ng–/– mice) demonstrate increased alcohol drinking compared to wild‐type mice, while also showing less tolerance to the effect of alcohol. To identify the molecular mechanism related to alcohol seeking, both in vivo microdialysis and label‐free quantification proteomics comparing Ng genotype and effects of alcohol treatment on the NAc are utilized. There is significant difference in glutamate and gamma‐aminobutyric acid (GABA) neurotransmission between genotypes; however, alcohol administration normalizes both glutamate and GABA levels in the NAc. Using label‐free proteomics, 427 protein expression changes are identified against alcohol treatment in the NAc among 4347 total proteins detected. Bioinformatics analyses reveal significant molecular differences in Ng null mice in response to acute alcohol treatment. Ingenuity pathway analysis found that the AKT network is altered significantly between genotypes, which may increase the sensitivity of alcohol in Ng null mice. The pharmacoproteomics results presented here illustrate a possible molecular basis of the alcohol sensitivity through Ng signaling in the NAc.

    更新日期:2019-12-23
  • Proteomics Analysis of Candida albicans dnm1 Haploid Mutant Unraveled the Association between Mitochondrial Fission and Antifungal Susceptibility
    Proteomics (IF 3.106) Pub Date : 2019-12-18
    Thuyen Truong; Guisheng Zeng; Teck Kwang Lim; Tong Cao; Li Mei Pang; Yew Mun Lee; Qingsong Lin; Yue Wang; Chaminda Jayampath Seneviratne

    Candida albicans is a major fungal pathogen, accounting for approximately 15% of healthcare infections with associated mortality as high as 40% in the case of systemic candidiasis. Antifungal agents for C. albicans infections are limited, and rising resistance is an inevitable problem. Therefore, understanding the mechanism behind antifungal responses is among the top research focuses in combating Candida infections. Herein, the recently developed C. albicans haploid model is employed to examine the association between mitochondrial fission, regulated by Dnm1, and the pathogen's response to antifungals. Proteomic analysis of dnm1Δ and its wild‐type haploid parent, GZY803, reveal changes in proteins associated with mitochondrial structures and functions, cell wall, and plasma membrane. Antifungal susceptibility testing revealed that dnm1Δ is more susceptible to SM21, a novel antifungal, than GZY803. Analyses of reactive oxygen species release, antioxidant response, lipid peroxidation, and membrane damages uncover an association between dnm1Δ and the susceptibility to SM21. Dynasore‐induced mitochondrial inhibition in SC5314 diploids corroborate the findings. Interestingly, Dynasore‐primed SC5314 cultures exhibit increased susceptibility to all antifungals tested. These data suggest an important contribution of mitochondrial fission in antifungal susceptibility of C. albicans. Hence, mitochondrial fission can be a potential target for combined therapy in anti‐C. albicans treatment.

    更新日期:2019-12-19
  • Scalable Data Analysis in Proteomics and Metabolomics Using BioContainers and Workflows Engines
    Proteomics (IF 3.106) Pub Date : 2019-12-18
    Yasset Perez‐Riverol; Pablo Moreno

    The recent improvements in mass spectrometry instruments and new analytical methods are increasing the intersection between proteomics and big data science. In addition, bioinformatics analysis is becoming increasingly complex and convoluted, involving multiple algorithms and tools. A wide variety of methods and software tools have been developed for computational proteomics and metabolomics during recent years, and this trend is likely to continue. However, most of the computational proteomics and metabolomics tools are designed as single‐tiered software application where the analytics tasks cannot be distributed, limiting the scalability and reproducibility of the data analysis. In this paper the key steps of metabolomics and proteomics data processing, including the main tools and software used to perform the data analysis, are summarized. The combination of software containers with workflows environments for large‐scale metabolomics and proteomics analysis is discussed. Finally, a new approach for reproducible and large‐scale data analysis based on BioContainers and two of the most popular workflow environments, Galaxy and Nextflow, is introduced to the proteomics and metabolomics communities.

    更新日期:2019-12-19
  • Proteomic Insights into Endometrial Receptivity and Embryo‐Endometrial Epithelium Interaction for Implantation; Critical Determinants of Fertility
    Proteomics (IF 3.106) Pub Date : 2019-12-11
    Jemma Evans, Jennifer Hutchison, Lois A. Salamonsen, David W. Greening

    In vitro fertilization has overcome infertility issues for many couples. However, achieving implantation of a viable embryo into the maternal endometrium remains a limiting step in optimizing pregnancy success. The molecular mechanisms which characterize the transient state of endometrial receptivity, critical in enabling embryo‐endometrial interactions, and proteins which underpin adhesion at the implantation interface, are limited in humans despite these temporally regulated processes fundamental to life. Hence, failure of implantation remains the ‘final frontier’ in infertility. We utilized a human co‐culture model utilizing spheroids of a trophectoderm (trophoblast stem) cell line, derived from pre‐implantation human embryos, and primary human endometrial epithelial cells, to functionally identify ‘fertile’ versus ’infertile’ endometrial epithelium based on adhesion between these cell types. Quantitative proteomics identified proteins associated with human endometrial epithelial receptivity (‘epithelial receptome’) and trophectoderm adhesion (‘adhesome’). As validation, key ‘epithelial receptome’ proteins (MAGT‐1/CDA/LGMN/KYNU/PC4) localized to the epithelium of receptive phase (mid‐secretory) endometrium obtained from fertile, normally cycling women but were largely absent from non‐receptive (proliferative) phase tissues. We demonstrate factors involved in embryo‐epithelium interaction in successive temporal stages of endometrial receptivity and implantation and provide potential targets for improving fertility, enhancing potential to become pregnant either naturally or in a clinical setting.

    更新日期:2019-12-11
  • Quantitative Proteomics Reveals Remodeling of Protein Repertoire Across Life Phases of Daphnia pulex
    Proteomics (IF 3.106) Pub Date : 2019-12-10
    Leonid Peshkin, Myriam Boukhali, Wilhelm Haas, Marc W. Kirschner, Lev Y. Yampolsky

    Although the microcrustacean Daphnia is becoming an organism of choice for proteomic studies, protein expression across its life cycle have not been fully characterized. Proteomes of adult females, juveniles, asexually produced embryos, and the ephippia‐resting stages containing sexually produced diapausing freezing‐ and desiccation‐resistant embryos are analyzed. Overall, proteins with known molecular functions are more likely to be detected than proteins with no detectable orthology. Similarly, proteins with stronger gene model support in two independent genome assemblies can be detected, than those without such support. This suggests that the proteomics pipeline can be applied to verify hypothesized proteins, even given questionable reference gene models. In particular, upregulation of vitellogenins and downregulation of actins and myosins in embryos of both types, relative to juveniles and adults, and overrepresentation of cell‐cycle related proteins in the developing embryos, relative to diapausing embryos and adults, are observed. Upregulation of small heat‐shock proteins and peroxidases, as well as overrepresentation of stress‐response proteins in the ephippium relative to the asexually produced non‐diapausing embryos, is found. The ephippium also shows upregulation of three trehalose‐synthesis proteins and downregulation of a trehalose hydrolase, consistent with the role of trehalose in protection against freezing and desiccation.

    更新日期:2019-12-11
  • Receptor‐Like Kinase Phosphorylation of Arabidopsis Heterotrimeric G‐Protein Gα ‐Subunit AtGPA1
    Proteomics (IF 3.106) Pub Date : 2019-12-10
    Haiyan Jia, Gaoyuan Song, Emily G. Werth, Justin W. Walley, Leslie M. Hicks, Alan M. Jones

    As molecular on–off switches, heterotrimeric G protein complexes, comprised of a Gα subunit and an obligate Gβγ dimer, transmit extracellular signals received by G protein–coupled receptors (GPCRs) to cytoplasmic targets that respond to biotic and abiotic stimuli. Signal transduction is modulated by phosphorylation of GPCRs and G protein complexes. In Arabidopsis thaliana, the Gα subunit AtGPA1 is phosphorylated by the receptor‐like kinase (RLK) BRI1‐associated Kinase 1 (BAK1), but the extent that other RLKs phosphorylates AtGPA1 is unknown. Twenty‐two trans‐phosphorylation sites on AtGPA1 are mapped by 12 RLKs hypothesized to act in the Arabidopsis G protein signaling pathway. Cis‐phosphorylation sites are also identified on these RLKs, some newly shown to be dual specific kinases. Multiple sites are present in the core AtGPA1 functional units, including pSer52 and/or pThr53 of the conserved P‐loop that directly binds nucleotide/phosphate, pThr164, and pSer175 from αE helix in the intramolecular domain interface for nucleotide exchange and GTP hydrolysis, and pThr193 and/or pThr194 in Switch I (SwI) that coordinates nucleotide exchange and protein partner binding. Several AtGPA1 S/T phosphorylation sites are potentially nucleotide‐dependent phosphorylation patterns, such as Ser52/Thr53 in the P‐loop and Thr193 and/or Thr194 in SwI.

    更新日期:2019-12-11
  • Reliability of Protein Abundance and Synthesis Measurements in Human Skeletal Muscle
    Proteomics (IF 3.106) Pub Date : 2019-12-06
    Kanchana Srisawat, Katie Hesketh, Matt Cocks, Juliette Strauss, Ben J. Edwards, Paulo J. Lisboa, Sam Shepherd, Jatin G. Burniston

    The repeatability of dynamic proteome profiling (DPP), which is a novel technique for measuring the relative abundance (ABD) and fractional synthesis rate (FSR) of proteins in humans, is investigated. LC–MS analysis is performed on muscle samples taken from male participants (n = 4) that consumed 4 × 50 mL doses of deuterium oxide (2H2O) per day for 14 days. ABD is measured by label‐free quantitation and FSR is calculated from time‐dependent changes in peptide mass isotopomer abundances. One‐hundred one proteins have at least one unique peptide and are used in the assessment of protein ABD. Fifty‐four of these proteins meet more stringent criteria and are used in the assessment of FSR data. The median (M), lower‐, (Q1) and upper‐quartile (Q3) values for protein FSR (%/d) are M = 1.63, Q1 = 1.07, and Q3 = 3.24, respectively. The technical CV of ABD data has a median value of 3.6% (Q1 1.7% to Q3 6.7%), whereas the median CV of FSR data is 10.1% (Q1 3.5% to Q3 16.5%). These values compare favorably against other assessments of technical repeatability of proteomics data, which often set a CV of 20% as the upper bound of acceptability.

    更新日期:2019-12-06
  • Nanomaterials in the Environment Acquire an “Eco‐Corona” Impacting their Toxicity to Daphnia Magna—a Call for Updating Toxicity Testing Policies
    Proteomics (IF 3.106) Pub Date : 2019-12-06
    Fatima Nasser, Julia Constantinou, Iseult Lynch

    Nanomaterials (NMs) are particles with at least one dimension between 1 and 100 nm and a large surface area to volume ratio, providing them with exceptional qualities that are exploited in a variety of industrial fields. Deposition of NMs into environmental waters during or after use leads to the adsorption of an ecological (eco‐) corona, whereby a layer of natural biomolecules coats the NM changing its stability, identity and ultimately toxicity. The eco‐corona is not currently incorporated into ecotoxicity tests, although it has been shown to alter the interactions of NMs with organisms such as Daphnia magna (D. magna). Here, the literature on environmental biomolecule interactions with NMs is synthesized and a framework for understanding the eco‐corona composition and its role in modulating NMs ecotoxicity is presented, utilizing D. magna as a model. The importance of including biomolecules as part of the current international efforts to update the standard testing protocols for NMs, is highlighted. Facilitating the formation of an eco‐corona prior to NMs ecotoxicity testing will ensure that signaling pathways perturbed by the NMs are real rather than being associated with the damage arising from reactive NM surfaces “acquiring” a corona by pulling biomolecules from the organism's surface.

    更新日期:2019-12-06
  • Sarcopenia: Tilting the Balance of Protein Homeostasis
    Proteomics (IF 3.106) Pub Date : 2019-12-04
    Kuan Ting Tan, Seok‐Ting Jamie Ang, Shih‐Yin Tsai

    Sarcopenia, defined as age‐associated decline of muscle mass and function, is a risk factor for mortality and disability, and comorbid with several chronic diseases such as type II diabetes and cardiovascular diseases. Clinical trials showed that nutritional supplements had positive effects on muscle mass, but not on muscle function and strength, demonstrating our limited understanding of the molecular events involved in the ageing muscle. Protein homeostasis, the equilibrium between protein synthesis and degradation, is proposed as the major mechanism underlying the development of sarcopenia. As the key central regulator of protein homeostasis, the mammalian target of rapamycin (mTOR) is proposed to be essential for muscle hypertrophy. Paradoxically, sustained activation of mTOR complex 1 (mTORC1) is associated with a loss of sensitivity to extracellular signaling in the elderly. It is not understood why sustained mTORC1 activity, which should induce muscle hypertrophy, instead results in muscle atrophy. Here, recent findings on the implications of disrupting protein homeostasis on muscle physiology and sarcopenia development in the context of mTOR/protein kinase B (AKT) signaling are reviewed. Understanding the role of these molecular mechanisms during the ageing process will contribute towards the development of targeted therapies that will improve protein metabolism and reduce sarcopenia.

    更新日期:2019-12-05
  • Profiling Differentially Abundant Proteins by Overexpression of Three Putative Methyltransferases in Xanthomonas axonopodis pv. glycines
    Proteomics (IF 3.106) Pub Date : 2019-12-02
    Hye‐Jee Park, Jongchan Lee, Minyoung Kim, Sang‐Wook Han

    Methyltransferases (MTases) are enzymes that modify specific substrates by adding a methyl group using S‐adenosyl‐l‐methionine. Functions of MTases have been extensively studied in eukaryotic organisms and animal pathogenic bacteria. Despite their importance, mechanisms underlying MTase function in plant pathogenic bacteria have not been studied in depth, as is the case of Xanthomonas axonopodis pv. glycines (Xag) that causes bacterial pustule disease in soybean crops worldwide. Here, the association between Xag proteome alterations and three MTase‐overexpressing strains, Xag(XgMT1), Xag(XgMT2), and Xag(XgMT3), compared to Xag carrying an empty vector, Xag(EV) is reported. Using label‐free shotgun comparative proteomic analysis, proteins are identified in all three biological replicates of the four strains and ranged from 1004 to 1082. In comparative analyses, 124, 135, and 134 proteins are differentially changed (over twofold) by overexpression of XgMT1, XgMT2, and XgMT3, respectively. These proteins are also categorized using cluster of orthologous group (COG) analyses, allowing postulation of biological mechanisms associated with three MTases in Xag. COGs reveal that the three MTases may play distinct roles, although some functions may overlap. These results are expected to allow new insight into understanding and predicting the biological functions of MTases in plant pathogenic bacteria. Data are available via ProteomeXchange (Identifier PXD012590).

    更新日期:2019-12-02
  • Deep Profiling of Cellular Heterogeneity by Emerging Single‐Cell Proteomic Technologies
    Proteomics (IF 3.106) Pub Date : 2019-12-02
    Liwei Yang, Justin George, Jun Wang

    The ability to comprehensively profile cellular heterogeneity in functional proteome is crucial in advancing the understanding of cell behavior, organism development, and disease mechanisms. Conventional bulk measurement by averaging the biological responses across a population often loses the information of cellular variations. Single‐cell proteomic technologies are becoming increasingly important to understand and discern cellular heterogeneity. The well‐established methods for single‐cell protein analysis based on flow cytometry and fluorescence microscopy are limited by the low multiplexing ability owing to the spectra overlap of fluorophores for labeling antibodies. Recent advances in mass spectrometry (MS), microchip, and reiterative staining‐based techniques for single‐cell proteomics have enabled the evaluation of cellular heterogeneity with high throughput, increased multiplexity, and improved sensitivity. In this review, the principles, developments, advantages, and limitations of these advanced technologies in analysis of single‐cell proteins, along with their biological applications to study cellular heterogeneity, are described. At last, the remaining challenges, possible strategies, and future opportunities that will facilitate the improvement and broad applications of single‐cell proteomic technologies in cell biology and medical research are discussed.

    更新日期:2019-12-02
  • Organismal Aging and Oxidants Beyond Macromolecules Damage
    Proteomics (IF 3.106) Pub Date : 2019-11-19
    Marie‐Veronique Clement, Le Luo

    The relationship between oxidants and organismal aging was first articulated through the free radical theory of aging. One of the major predictions of the free radical theory of aging is that oxidative stress shortens organisms’ lifespan because of an increased level of oxidants, which are potentially damaging to macromolecules. However, challenging the role of oxidants in age‐related diseases, there is now sufficient evidence that anti‐oxidant supplements do not provide any significant health benefits. Interestingly, in addition to an increase in oxidant‐mediated macromolecules damage, there is convincing experimental data to support the role for senescent cells in the process of aging. Here, we review the current knowledge regarding the role of oxidants and cellular senescence in organismal aging and propose that, in addition to the role of oxidants as inducers of macromolecular damage, oxidants might also function as regulators of signalling pathways involved in the establishment of cellular senescence. If this role for oxidants is established, it might be necessary to modify the free radical theory of aging from “Organisms age because cells accumulate reactive oxygen species‐dependent damage over time” to: “Organisms age because cells accumulate oxidants’‐dependent damage and oxidants’‐dependent senescent characteristics over time”.

    更新日期:2019-11-20
  • Magnetic particles as Powerful Purification Tool for High Sensitive Mass Spectrometric Screening Procedures
    Proteomics (IF 3.106) Pub Date : 2010-08-19
    Jochen F. Peter, Angela M. Otto, Bernhard Wolf

    The effective isolation and purification of proteins from biological fluids is the most crucial step for a successful protein analysis when only minute amounts are available. While conventional purification methods like dialysis, ultrafiltration or protein precipitation often leads to a marked loss of protein, solid phase extraction (SPE) with small‐sized particles is a powerful alternative. The implementation of particles with superparamagnetic cores facilitates the handling of those particles and allows the application of particles in the nm‐ to low ìm range.

    更新日期:2019-11-18
  • Targeted and Untargeted Proteomics Approaches in Biomarker Development
    Proteomics (IF 3.106) Pub Date : 2019-11-15
    Constance A. Sobsey, Sahar Ibrahim, Vincent R. Richard, Vanessa Gaspar, Georgia Mitsa, Vincent Lacasse, René P. Zahedi, Gerald Batist, Christoph H. Borchers

    An enormous amount of research effort has been devoted to biomarker discovery and validation. With the completion of the human genome, proteomics is now playing an increasing role in this search for new and better biomarkers. Here, we review what leads to successful biomarker development and consider how these features may be applied in the context of proteomic biomarker research. The “fit‐for‐purpose” approach to biomarker development suggests that untargeted proteomic approaches may be better suited for early stages of biomarker discovery, while targeted approaches are preferred for validation and implementation. A systematic screening of published biomarker articles using mass spectrometry‐based proteomics reveals that while both targeted and untargeted technologies are used in proteomic biomarker development, most researchers do not combine these approaches. We discuss (i) the reasons for this discrepancy, (ii) how proteomic technologies can overcome technical challenges that seem to limit their translation into the clinic, and (iii) how mass spectrometry can improve, complement, or replace existing clinically‐important assays in the future.

    更新日期:2019-11-15
  • Thermoregulation in the Aging Population and Practical Strategies to Overcome a Warmer Tomorrow
    Proteomics (IF 3.106) Pub Date : 2019-11-14
    Chee Chong Shawn Tan, Li Kang Karen Chin, Ivan Cherh Chiet Low

    As global temperatures continue to rise, improving thermal tolerance in the aged population is crucial to counteract age‐associated impairments in thermoregulatory function. Impairments in reflex cutaneous vasodilation and sweating response can augment the vulnerability of older adults to heat‐related injuries following exposure to heat stress. Mechanisms underlying a compromised cutaneous vasodilation are suggested to include reduced sympathetic neural drive, diminished cholinergic co‐transmitter contribution, and altered second messenger signaling events. On the other hand, impairments in sweating response are ascribed to reduced sweat gland cholinergic sensitivity and altered cyclooxygenase and nitric oxide signaling. Several practical mitigation strategies such as exercise, passive heating, and behavioral adaptations are proposed as means to overcome heat stress and improve thermal tolerance in the aged. Aerobic exercise training is shown to be amongst the most effective ways to enhance thermoregulatory function. However, in elderly with limited exercise capability due to chronic diseases and mobility issues, passive heating can serve as a functional alternative as it has been shown to confer similar benefits to that of exercise training. Supplementary to exercise training and passive heating, behavioral adaptations can be applied to further enhance the heat‐preparedness of the aged.

    更新日期:2019-11-14
  • Adenosine‐to‐Inosine RNA Editing in Mouse and Human Brain Proteomes
    Proteomics (IF 3.106) Pub Date : 2019-11-13
    Lev I. Levitsky, Anna A. Kliuchnikova, Ksenia G. Kuznetsova, Dmitry S. Karpov, Mark V. Ivanov, Mikhail A. Pyatnitskiy, Olga V. Kalinina, Mikhail V. Gorshkov, Sergei A. Moshkovskii

    Proteogenomics is based on the use of customized genome or RNA sequencing databases for interrogation of shotgun proteomics data in search for proteome‐level evidence of genome variations or RNA editing. In this work, the products of adenosine‐to‐inosine RNA editing in human and murine brain proteomes are identified using publicly available brain proteome LC‐MS/MS datasets and an RNA editome database compiled from several sources. After filtering of false‐positive results, 20 and 37 sites of editing in proteins belonging to 14 and 32 genes are identified for murine and human brain proteomes, respectively. Eight sites of editing identified with high spectral counts overlapped between human and mouse brain samples. Some of these sites have been previously reported using orthogonal methods, such as α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) glutamate receptors, CYFIP2, coatomer alpha. Also, differential editing between neurons and microglia is demonstrated in this work for some of the proteins from primary murine brain cell cultures. Because many edited sites are still not characterized functionally at the protein level, the results provide a necessary background for their further analysis in normal and diseased cells and tissues using targeted proteomic approaches.

    更新日期:2019-11-13
  • Interspecies Comparison of the Bacterial Response to Allicin Reveals Species‐Specific Defense Strategies
    Proteomics (IF 3.106) Pub Date : 2019-11-12
    Dominik Wüllner, Annika Haupt, Pascal Prochnow, Roman Leontiev, Alan J. Slusarenko, Julia E. Bandow

    Allicin, a broad‐spectrum antimicrobial agent from garlic, disrupts thiol and redox homeostasis, proteostasis, and cell membrane integrity. Since medicine demands antimicrobials with so far unexploited mechanisms, allicin is a promising lead structure. While progress is being made in unraveling its mode of action, little is known on bacterial adaptation strategies. Some isolates of Pseudomonas aeruginosa and Escherichia coli withstand exposure to high allicin concentrations due to as yet unknown mechanisms. To elucidate resistance and sensitivity‐conferring cellular processes, the acute proteomic responses of a resistant P. aeruginosa strain and the sensitive species Bacillus subtilis are compared to the published proteomic response of E. coli to allicin treatment. The cellular defense strategies share functional features: proteins involved in translation and maintenance of protein quality, redox homeostasis, and cell envelope modification are upregulated. In both Gram‐negative species, protein synthesis of the majority of proteins is downregulated while the Gram‐positive B. subtilis responded by upregulation of multiple regulons. A comparison of the B. subtilis proteomic response to a library of responses to antibiotic treatment reveals 30 proteins specifically upregulated by allicin. Upregulated oxidative stress proteins are shared with nitrofurantoin and diamide. Microscopy‐based assays further indicate that in B. subtilis cell wall integrity is impaired.

    更新日期:2019-11-13
  • Proteomic Analysis of the Lake Trout (Salvelinus namaycush) Liver Identifies Proteins from Evolutionarily Close and ‐Distant Fish Relatives
    Proteomics (IF 3.106) Pub Date : 2019-11-12
    Emmalyn J. Dupree, Bernard S. Crimmins, Thomas M. Holsen, Costel C. Darie

    Lake trout are used as bioindicators for toxics exposure in the Great Lakes ecosystem. Here the first lake trout (Salvelinus namaycush) liver proteomics study is performed and searched against specific databases: (NCBI and UniProtKB) Salvelinus, Salmonidae, Actinopterygii, and Oncorhynchus mykiss, and the more distant relative, Danio rerio. In the biological replicate 1 (BR1), technical replicate 1 (TR1), (BR1TR1), a large number of lake trout liver proteins are not in the Salvelinus protein database, suggesting that lake trout liver proteins have homology to some proteins from the Salmonidae family and Actinopterygii class, and to Oncorhynchus mykiss and Danio rerio, two more highly studied fish. In the NCBI search, 4194 proteins are identified: 3069 proteins in Actinopterygii, 1617 in Salmonidae, 68 in Salvelinus, 568 in Oncorhynchus mykiss, and 946 in Danio rerio protein databases. Similar results are observed in the UniProtKB searches of BR1RT1, as well as in a technical replicate (BR1TR2), and then in a second biological replicate experiment, with two technical replicates (BR2TR1 and BR2TR2). This study opens the possibility of identifying evolutionary relationships (i.e., adaptive mutations) between various groups (i.e., zebrafish, rainbow trout, Salmonidae, Salvelinus and lake trout) through evolutionary proteomics. Data are available via the PRIDE Q2 (PXD011924).

    更新日期:2019-11-13
  • Mass Spectrometry‐Based Analysis of Time‐Resolved Proteome Quantification
    Proteomics (IF 3.106) Pub Date : 2019-11-11
    Alberto Valdés, Sara Bergström Lind

    The aspect of time is essential in biological processes and thus it is important to be able to monitor signaling molecules through time. Proteins are key players in cellular signaling and they respond to many stimuli and change their expression in many time‐dependent processes. Mass spectrometry (MS) is an important tool for studying proteins, including their posttranslational modifications and their interaction partners—both in qualitative and quantitative ways. In order to distinguish the different trends over time, proteins, modification sites, and interacting proteins must be compared between different time points, and therefore relative quantification is preferred. In this review, the progress and challenges for MS‐based analysis of time‐resolved proteome dynamics are discussed. Further, aspects on model systems, technologies, sampling frequencies, and presentation of the dynamic data are discussed.

    更新日期:2019-11-11
  • MALDI–MS Profiling to Address Honey Bee Health Status under Bacterial Challenge through Computational Modeling
    Proteomics (IF 3.106) Pub Date : 2019-11-11
    Karim Arafah, Sébastien Nicolas Voisin, Victor Masson, Cédric Alaux, Yves Le Conte, Michel Bocquet, Philippe Bulet

    Honey bees play a critical role in the maintenance of plant biodiversity and sustainability of food webs. In the past few decades, bees have been subjected to biotic and abiotic threats causing various colony disorders. Therefore, monitoring solutions to help beekeepers to improve bee health are necessary. Matrix‐assisted laser desorption ionization–mass spectrometry (MALDI–MS) profiling has emerged within this decade as a powerful tool to identify in routine micro‐organisms and is currently used in real‐time clinical diagnosis. MALDI BeeTyping is developed to monitor significant hemolymph molecular changes in honey bees upon infection with a series of entomopathogenic Gram‐positive and ‐negative bacteria. A Serratia marcescens strain isolated from one naturally infected honey bee collected from the field is also considered. A series of hemolymph molecular mass fingerprints is individually recorded and to the authors' knowledge, the first computational model harboring a predictive score of 97.92% and made of nine molecular signatures that discriminate and classify the honey bees’ systemic response to the bacteria is built. Hence, the model is challenged by classifying a training set of hemolymphs and an overall recognition of 91.93% is obtained. Through this work, a novel, time and cost saving high‐throughput strategy that addresses honey bee health on an individual scale is introduced.

    更新日期:2019-11-11
  • motifeR: An Integrated Web Software for Identification and Visualization of Protein Posttranslational Modification Motifs
    Proteomics (IF 3.106) Pub Date : 2019-11-07
    Shisheng Wang, Yue Cai, Jingqiu Cheng, Wenxue Li, Yansheng Liu, Hao Yang

    With an exponential growth in applications identifying protein post‐translational modifications via mass spectrometry, discovery and presentation of motifs surrounding those modification sites have become increasingly desirable. Despite a few tools being designed, there is still a scarcity of effective and polyfunctional software for such analysis and illustrations. In this study, a versatile and user‐friendly web tool is developed, motifeR, for extracting and visualizing statistically significant motifs from large datasets. Particularly, several functions are also integrated for processing multi‐modification sites enrichment. Public datasets are applied to test their usability, indicating that some concurrent modification sites may form motifs and that peptides with low location probability may be not identified randomly and can be included to support motif discovery. In addition, for human phosphoproteomics datasets, the characterization of differential kinase signaling networks can be estimated and modeled by combining kinase‐substrate relations based on the NetworKIN database as an optional feature for users. The motifeR toolkit can be conveniently operated by any scientific community or individuals, even those without any bioinformatics background and is freely available at https://www.omicsolution.org/wukong/motifeR.

    更新日期:2019-11-07
  • Trajectories of Aging: How Systems Biology in Yeast Can Illuminate Mechanisms of Personalized Aging
    Proteomics (IF 3.106) Pub Date : 2019-11-04
    Matthew M. Crane, Kenneth L. Chen, Ben W. Blue, Matt Kaeberlein

    All organisms age, but the extent to which all organisms age the same way remains a fundamental unanswered question in biology. Across species, it is now clear that at least some aspects of aging are highly conserved and are perhaps universal, but other mechanisms of aging are private to individual species or sets of closely related species. Within the same species, however, it has generally been assumed that the molecular mechanisms of aging are largely invariant from one individual to the next. With the development of new tools for studying aging at the individual cell level in budding yeast, recent data has called this assumption into question. There is emerging evidence that individual yeast mother cells may undergo fundamentally different trajectories of aging. Individual trajectories of aging are difficult to study by traditional population level assays, but through the application of systems biology approaches combined with novel microfluidic technologies, it is now possible to observe and study these phenomena in real time. Understanding the spectrum of mechanisms that determine how different individuals age is a necessary step toward the goal of personalized geroscience, where healthy longevity is optimized for each individual.

    更新日期:2019-11-04
  • 更新日期:2019-11-01
  • Proteome Profile and Genome Refinement of the Tomato-Pathogenic Bacterium Clavibacter michiganensis subsp. michiganensis.
    Proteomics (IF 3.106) Pub Date : 2019-01-17
    F Christopher Peritore-Galve,David J Schneider,Yong Yang,Theodore W Thannhauser,Christine D Smart,Paul Stodghill

    更新日期:2019-11-01
  • Comparative proteomics analysis reveals roles for FADD in the regulation of energy metabolism and proteolysis pathway in mouse embryonic fibroblast.
    Proteomics (IF 3.106) Pub Date : 2013-06-08
    Hongqin Zhuang,Ziyi Gan,Weiwei Jiang,Xiangyu Zhang,Zi-Chun Hua

    Fas-associated death domain-containing protein (FADD) is a classical apoptotic pathway adaptor. Further studies revealed that it also plays essential roles in nonapoptotic processes, which is assumed to be regulated by its phosphorylation. However, the exact mechanisms are still poorly understood. To study the nonapoptotic effects of FADD, a comprehensive strategy of proteomics identification combined with bioinformatic analysis was undertaken to identify proteins differentially expressed in three cell lines containing FADD and its mutant, FADD-A and FADD-D. The cell lines were thought to bear wild-type FADD, unphosphorylated FADD mimic and constitutive phosphorylated FADD mimic, respectively. A total of 47 proteins were identified to be significantly changed due to FADD phosphorylation. Network analysis using MetaCore™ identified a number of changed proteins that were involved in cellular metabolic process, including lipid metabolism, fatty acid metabolism, glycolysis, and oxidative phosphorylation. The finding that FADD-D cell line showed an increase in fatty acid oxidation argues that it could contribute to the leaner phenotype of FADD-D mice as reported previously. In addition, six proteins related to the ubiquitin-proteasome pathway were also specifically overexpressed in FADD-D cell line. Finally, the c-Myc gene represents a convergent hub lying at the center of dysregulated pathways, and was upregulated in FADD-D cells. Taken together, these studies allowed us to conclude that impaired mitochondrial function and proteolysis might play pivotal roles in the dysfunction associated with FADD phosphorylation-induced disorders.

    更新日期:2019-11-01
  • Standard abbreviations.
    Proteomics (IF 3.106) Pub Date : 2018-06-07

    更新日期:2019-11-01
  • Part II: Special Issue on Extracellular Vesicles and Exosomes.
    Proteomics (IF 3.106) Pub Date : 2019-04-17
    Richard J Simpson,David W Greening

    更新日期:2019-11-01
  • 更新日期:2019-11-01
  • ProSight Lite: graphical software to analyze top-down mass spectrometry data.
    Proteomics (IF 3.106) Pub Date : 2015-04-02
    Ryan T Fellers,Joseph B Greer,Bryan P Early,Xiang Yu,Richard D LeDuc,Neil L Kelleher,Paul M Thomas

    Many top-down proteomics experiments focus on identifying and localizing PTMs and other potential sources of “mass shift” on a known protein sequence. A simple application to match ion masses and facilitate the iterative hypothesis testing of PTM presence and location would assist with the data analysis in these experiments. ProSight Lite is a free software tool for matching a single candidate sequence against a set of mass spectrometric observations. Fixed or variable modifications, including both PTMs and a select number of glycosylations, can be applied to the amino acid sequence. The application reports multiple scores and a matching fragment list. Fragmentation maps can be exported for publication in either portable network graphic (PNG) or scalable vector graphic (SVG) format. ProSight Lite can be freely downloaded from http://prosightlite.northwestern.edu, installs and updates from the web, and requires Windows 7 or a higher version.

    更新日期:2019-11-01
  • Proteomics in the fruit tree science arena: new insights into fruit defense, development, and ripening.
    Proteomics (IF 3.106) Pub Date : 2013-08-30
    Athanassios Molassiotis,Georgia Tanou,Panagiota Filippou,Vasileios Fotopoulos

    Fruit tree crops are agricultural commodities of high economic importance, while fruits also represent one of the most vital components of the human diet. Therefore, a great effort has been made to understand the molecular mechanisms covering fundamental biological processes in fruit tree physiology and fruit biology. Thanks to the development of cutting-edge "omics" technologies such as proteomic analysis, scientists now have powerful tools to support traditional fruit tree research. Such proteomic analyses are establishing high-density 2DE reference maps and peptide mass fingerprint databases that can lead fruit science into a new postgenomic research era. Here, an overview of the application of proteomics in key aspects of fruit tree physiology as well as in fruit biology, including defense responses to abiotic and biotic stress factors, is presented. A panoramic view of ripening-related proteins is also discussed, as an example of proteomic application in fruit science.

    更新日期:2019-11-01
  • Proteomic insights into seed germination in response to environmental factors.
    Proteomics (IF 3.106) Pub Date : 2013-08-30
    Longyan Tan,Sixue Chen,Tai Wang,Shaojun Dai

    Seed germination is a critical process in the life cycle of higher plants. During germination, the imbibed mature seed is highly sensitive to different environmental factors.However, knowledge about the molecular and physiological mechanisms underlying the environmental effects on germination has been lacking. Recent proteomic work has provided invaluable insight into the molecular processes in germinating seeds of Arabidopsis, rice (Oryza sativa), soybean (Glycine max), barley (Hordeum vulgare), maize (Zeamays), tea (Camellia sinensis), European beech (Fagus sylvatica), and Norway maple (Acer platanoides) under different treatments including metal ions (e.g. copper and cadmium), drought, low temperature, hormones, and chemicals (gibberellic acid, abscisic acid, salicylic acid, and α-amanitin), as well as Fusarium graminearum infection. A total of 561 environmental factor-responsive proteins have been identified with various expression patterns in germinating seeds. The data highlight diverse regulatory and metabolic mechanisms upon seed germination, including induction of environmental factor-responsive signaling pathways, seed storage reserve mobilization and utilization, enhancement of DNA repair and modification, regulation of gene expression and protein synthesis, modulation of cell structure, and cell defense. In this review, we summarize the interesting findings and discuss the relevance and significance for our understanding of environmental regulation of seed germination.

    更新日期:2019-11-01
  • Standard abbreviations.
    Proteomics (IF 3.106) Pub Date : null

    更新日期:2019-11-01
  • Cancer Omics: Adding Understanding to Knowledge.
    Proteomics (IF 3.106) Pub Date : 2019-09-27
    Hubert Hondermarck

    更新日期:2019-11-01
  • Analysis of the cerebellar proteome in a transgenic mouse model of inherited prion disease reveals preclinical alteration of calcineurin activity.
    Proteomics (IF 3.106) Pub Date : 2006-03-31
    Emiliano Biasini,Tania Massignan,Luana Fioriti,Valentina Rossi,Sara Dossena,Mario Salmona,Gianluigi Forloni,Valentina Bonetto,Roberto Chiesa

    Inherited prion diseases are linked to insertional and point mutations in the prion protein (PrP) gene, which favor conversion of PrP into a conformationally altered, pathogenic isoform. The cellular mechanism by which this process causes neurological dysfunction is unknown. Transgenic (Tg) (PG14) mice express a mouse PrP homolog of a nine-octapeptide insertion associated with an inherited prion disorder. These mice develop a progressive neurological syndrome characterized by ataxia and cerebellar atrophy due to synaptic degeneration in the molecular layer and massive apoptosis of granule neurons. To investigate the molecular events that may contribute to neurological dysfunction, we carried out a differential proteomic analysis of cerebella from Tg(PG14) mice at the preclinical, onset, and symptomatic phases of their neurological illness. 2-D maps of cerebellar proteins from Tg(PG14) mice were compared to those obtained from age-matched Tg(WT) mice that express wild-type PrP and remain healthy. Proteins whose levels were significantly modified in at least one stage of the Tg(PG14) disease were identified by PMF. Analysis detected a preclinical decrease of the calcium/calmodulin-dependent phosphatase calcineurin (CaN) in granule neurons, suggesting that dysregulation of CaN activity induced by mutant PrP may be responsible for the cerebellar dysfunction in Tg(PG14) mice.

    更新日期:2019-11-01
  • Transcriptomic and proteomic responses of human renal HEK293 cells to uranium toxicity.
    Proteomics (IF 3.106) Pub Date : 2005-01-27
    Odette Prat,Frédéric Berenguer,Véronique Malard,Emmanuelle Tavan,Nicole Sage,Gérard Steinmetz,Eric Quemeneur

    The industrial use of uranium, in particular depleted uranium, has pin-pointed the need to review its chemical impact on human health. Global methodologies, applied to the field of toxicology, have demonstrated their applicability to investigation of fine molecular mechanisms. This report illustrate the power of toxicogenomics to evaluate the involvement of certain genes or proteins in response to uranium. We particularly show that 25% of modulated genes concern signal transduction and trafficking, that the calcium pathway is heavily disturbed and that nephroblastomas-related genes are involved (WIT-1, STMN1, and STMN2). A set of 18 genes was deregulated whatever the concentration of toxicant, which could constitute a signature of uranium exposure. Moreover, a group of downregulated genes, with corresponding disappearing proteins (HSP90, 14-3-3 protein, HMGB1) in two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), are good candidates for use as biomarkers of uranium effects. These results reveal a cross-checking between transcriptomic and proteomic technologies. Moreover, our temporal gene expression profiles suggest the existence of a concentration threshold between adaptive response and severe cell deregulation. Our results confirm the involvement of genes already described and also provide new highlights on cellular response to uranium.

    更新日期:2019-11-01
  • The impact of blood contamination on the proteome of cerebrospinal fluid.
    Proteomics (IF 3.106) Pub Date : 2005-01-27
    Jin-Sam You,Valentina Gelfanova,Michael D Knierman,Frank A Witzmann,Mu Wang,John E Hale

    Human cerebrospinal fluid (CSF) is in direct contact with the brain extracellular space. Beside the secretion of CSF by the choroid plexus the fluid also derives directly from the brain by the ependymal lining of the ventricular system and the glial membrane and from blood vessels in the arachnoid. Therefore, biochemical change in the brain may be reflected in the CSF. CSF is a potential source of protein molecular indices of central nervous system function and pathology. However, various amounts of blood contamination in CSF may arise during sample acquisition. The concentration of protein in the CSF is only 0.2 to 0.5% that of blood. Minor contamination of CSF with blood during collection of the fluid may dramatically alter the protein profile confounding the identification of potential biomarkers. We have analyzed CSF and CSF spiked with increasing amounts of whole blood using proteomic techniques. We detected at least four blood specific highly abundant proteins: hemoglobin, catalase, peroxiredoxin and carbonic anhydrase I. These proteins can be used as blood contamination markers for proteomic analysis of CSF. Proteins in blood contaminated CSF samples were less stable compared to neat CSF at 37 degrees C suggesting that blood borne protease may induce protein degradation in CSF during sample acquisition. This analysis was aimed at identification of proteins found primarily in CSF, those found primarily in blood and assessment of the impact of blood contamination on those proteins found in both fluids.

    更新日期:2019-11-01
  • Proteomic analysis of mitochondria from Caenorhabditis elegans.
    Proteomics (IF 3.106) Pub Date : 2009-08-12
    Jing Li,Tanxi Cai,Peng Wu,Ziyou Cui,Xiulan Chen,Junjie Hou,Zhensheng Xie,Peng Xue,Linan Shi,Pingsheng Liu,John R Yates,Fuquan Yang

    Mitochondria play essential roles in cell physiological processes including energy production, metabolism, ion homeostasis, cell growth, aging and apoptosis. Proteomic strategies have been applied to the study of mitochondria since 1998; these studies have yielded decisive information about the diverse physiological functions of the organelle. As an ideal model biological system, the nematode Caenorhabditis elegans has been widely used in the study of several diseases, such as metabolic diseases and cancer. However, the mitochondrial proteome of C. elegans remains elusive. In this study, we purified mitochondria from C. elegans and performed a comprehensive proteomic analysis using the shotgun proteomic approach. A total of 1117 proteins have been identified with at least two unique peptides. Their physicochemical and functional characteristics, subcellular locations, related biological processes, and associations with human diseases, especially Parkinson's disease, are discussed. An orthology comparison was also performed between C. elegans and four other model organisms for a general depiction of the conservation of mitochondrial proteins during evolution. This study will provide new clues for understanding the role of mitochondria in the physiological and pathological processes of C. elegans.

    更新日期:2019-11-01
  • Strategies for shotgun identification of integral membrane proteins by tandem mass spectrometry.
    Proteomics (IF 3.106) Pub Date : 2008-09-10
    Bingwen Lu,Daniel B McClatchy,Jin Young Kim,John R Yates

    Integral membrane proteins (IMPs) are difficult to identify, mainly for two reasons: the hydrophobicity of IMPs and their low abundance. Sample preparation is a key component in the large-scale identification of IMPs. In this review, we survey strategies for shotgun identification of IMPs by MS/MS. We will discuss enrichment, solubilization, separation, and digestion of IMPs, and data analysis for membrane proteomics.

    更新日期:2019-11-01
  • Characterisation by proteomics of peribacteroid space and peribacteroid membrane preparations from pea (Pisum sativum) symbiosomes.
    Proteomics (IF 3.106) Pub Date : 2002-03-29
    Gerhard Saalbach,Pinar Erik,Stefanie Wienkoop

    The legume Rhizobium symbiosis leads to the formation of a new compartment in the plant cell, the symbiosome. This compartment harbours the bacteroids surrounded by a peribacteroid membrane (PBM) originating from the plant plasma membrane. The PBM and the space between the PBM and the bacteroid membrane, called peribacteroid space (PS), mediate the exchange of metabolites between the symbionts. Proteome analysis was used as an approach to characterise the proteins in the PBM and the PS. A standard differential centrifugation procedure including a Percoll gradient was used for symbiosome isolation from pea root nodules. Proteins in the PBM and PS fractions obtained from the symbiosomes were separated by two-dimensional gel electrophoresis, and 89 spots were analysed by tandem mass spectrometry. The proteins of 46 spots could be identified by database search. The results showed that PS and even PBM preparations from pea symbiosomes always contain abundant amounts of bacteroid proteins as a contaminate. Interestingly, in addition to a few PS/PBM proteins a number of endomembrane proteins (less likely representing a contaminate), including V-ATPase, BIP, and an integral membrane protein known from COPI-coated vesicles, were found in the PBM fraction, supporting the role of the endomembrane system in PBM biogenesis.

    更新日期:2019-11-01
  • Immunoproteomics of Helicobacter pylori infection and relation to gastric disease.
    Proteomics (IF 3.106) Pub Date : 2002-03-29
    Gaby Haas,Galip Karaali,Karl Ebermayer,Wolfram G Metzger,Stephanie Lamer,Ursula Zimny-Arndt,Susanne Diescher,Ulf B Goebel,Konstanze Vogt,Artur B Roznowski,Bertram J Wiedenmann,Thomas F Meyer,Toni Aebischer,Peter R Jungblut

    The Gram negative bacterium Helicobacter pylori is a human pathogen which infects the gastric mucosa and causes an inflammatory process leading to gastritis, ulceration and cancer. A systematic, proteome based approach was chosen to detect candidate antigens of H. pylori for diagnosis, therapy and vaccine development and to investigate potential associations between specific immune responses and manifestations of disease. Sera from patients with active H. pylori infection (n = 24), a control group with unrelated gastric disorders (n = 12) and from patients with gastric cancer (n = 6) were collected and analyzed for the reactivity against proteins of the strain HP 26695 separated by two-dimensional electrophoresis. Overall, 310 antigenic protein species were recognized by H. pylori positive sera representing about 17% of all spots separated. Out of the 32 antigens most frequently recognized by H. pylori positive sera, nine were newly identified and 23 were confirmed from other studies. Three newly identified antigens which belong to the 150 most abundant protein species of H. pylori, were specifically recognized by H. pylori positive sera: the predicted coding region HP0231, serine protease HtrA (HP1019) and Cag3 (HP0522). Other antigens were recognized differently by sera from gastritis and ulcer patients, which may identify them as candidate indicators for clinical manifestations. The data from these immunoproteomic analyses are added to our public database (http://www.mpiib-berlin.mpg.de/2D-PAGE). This platform enables one to compile many protein profiles and to integrate data from other studies, an approach which will greatly assist the search for more immunogenic proteins for diagnostic assays and vaccine design.

    更新日期:2019-11-01
  • Proteomic analysis of differential protein expression in human nasopharyngeal carcinoma cells induced by NAG7 transfection.
    Proteomics (IF 3.106) Pub Date : 2002-03-29
    Chen Tan,Jiang Li,Jieru Wang,Qiu Xiang,Xiaomei Zhang,Li Dong,Shourong Shen,Songping Liang,Guiyuan Li

    Nasopharyngeal carcinoma (NPC) is a commonly occurring tumor in southern China and south east Asia. A genetic factor has now been recognized to be associated with this cancer. A new gene, named NAG7, was cloned from the common minimal deletion region in 3p25.3-26.3. In order to investigate the function of NAG7 gene, proteomic methods were used to find and identify the differential proteins and expected to elucidate the mechanism of NAG7. The NAG7 eukaryotic expression vector was constructed and transfected into NPC cell line HNE1 with liposome. Twenty-two differential protein spots in transfected cells were found significant and reproducible using high-resolution two-dimensional electrophoresis. Nine proteins that were up-regulated and seven proteins that were down-regulated were identified by matrix assisted laser desorption/ionization time of flight mass spectrometry and database analysis. These proteins included growth arrest specific protein, DNA binding protein, caspase 6, pinch protein and ras-related protein rab-36, which are involved in cell cycling, transcription regulation, signaling pathways and apoptosis. NAG7 may exert its functions by mediating differential expression of these proteins.

    更新日期:2019-11-01
  • Towards complete analysis of the platelet proteome.
    Proteomics (IF 3.106) Pub Date : 2002-03-29
    Eric E O'Neill,Chris J Brock,Alexander F von Kriegsheim,Andrew C Pearce,Raymond A Dwek,Steve P Watson,Holger F Hebestreit

    Platelets exert a crucial function in haemostasis, wound repair, and the formation of vascular plugs, underlying thrombotic diseases such as stroke and myocardial infarction. Analysis of platelet biochemistry is largely dependent on protein analysis as platelets are anucleated cells providing little analytical target for DNA or RNA based strategies. Here we present data from our analysis of the human platelet proteome, the entire set of proteins building a platelet at a given point in time. Proteins were separated by two-dimensional electrophoresis (2-DE) using broad and narrow range pH gradients in the isoelectric focusing step. Consequently, a high-resolution 2-DE proteome map has been generated that comprises approximately 2300 different protein features. From the 536 protein features detected in the 4-5 pI range 284 features were identified by electrospray ionisation time of flight tandem mass spectrometry. These 284 proteins originate from 123 different open reading frames. This includes the five human proteins KIAA0193, KIAA0573, KIAA0830, WUGSC:H_DJ0777O23 protein, and cytokine receptor related protein 4, all isolated for the first time. The data are discussed with regard to proteome characteristics, protein function, and the high prevalence of signalling molecules. This study contributes to a more thorough and holistic understanding of platelet biology, helping to build the basis for future identification of new drug targets and therapeutic strategies.

    更新日期:2019-11-01
  • Primary skin fibroblasts as human model system for proteome analysis.
    Proteomics (IF 3.106) Pub Date : 2002-03-29
    Stefan Lehr,Jorg Kotzka,Birgit Knebel,Martina Schiller,Wilhelm Krone,Dirk Muller-Wieland

    Elucidation of cellular processes and their changes at the level of protein expression and post-translational modification patterns may allow identification of novel proteins and thereby mechanisms involved in the pathogenesis of multigenic diseases. The aim of this study was to test cultured, nontransformed primary fibroblasts derived from human skin biopsies as a suitable model system for proteome analysis. Therefore soluble protein fractions were separated on several overlapping ultrazoom gels covering the pH range from 3.5-9. Correlation analysis of gel-pairs revealed a highly reproducible protein expression pattern within (intra-assay) and between (inter-assay) independent experiments of a single fibroblast cell line (intra-cell line comparison). Spot intensity variations were less than a factor of two for more than 80% of identical spots. In addition, inter-cell line comparison exhibits no significant variations in spot intensities. To achieve further improvements in reproducibility we generated master gels for each pH range by combining averaged spot information derived from two different cell lines each analysed by two independent experiments using the raw master gel algorithm of the Z3 image analysis software. The resulting reference images of primary human fibroblasts provided a basis for investigating regulation by extracellular stimuli and drugs as well as their alterations in patients with different diseases.

    更新日期:2019-11-01
  • A strategy for mapping and neutralizing conformational immunogenic sites on protein therapeutics.
    Proteomics (IF 3.106) Pub Date : 2002-03-29
    Daniel I R Spencer,Lynda Robson,Des Purdy,Nick R Whitelegg,N Paul Michael,Jeetendra Bhatia,Surinder K Sharma,Anthony R Rees,Nigel P Minton,Richard H J Begent,Kerry A Chester

    Antibodies are highly specific recognition molecules which are increasingly being applied to target therapy in patients. One type of developmental antibody-based therapy is antibody directed enzyme prodrug therapy (ADEPT) for the treatment of cancer. In ADEPT, an antibody specific to a tumor marker protein delivers a drug-activating enzyme to the cancer. Subsequent intravenous administration of an inactive prodrug results in drug activation and cytotoxicity only within the locale of the tumor. Pilot clinical trials with chemical conjugates of the prodrug activating enzyme carboxypeptidase G2 (CPG2) chemically conjugated with an antibody to and carcinoembryonic antigen (CEA), have shown that CPG2-mediated ADEPT is effective but limited by formation of human antibodies to CPG2 (HACA). We have developed a recombinant fusion protein (termed MFE-CP) of CPG2 with an anti-CEA single chain Fv antibody fragment and we have developed methods to address the immunogenicity of this therapeutic. A HACA-reactive discontinuous epitope on MFE-CP was identified using the crystal structure of CPG2, filamentous phage technology and surface enhanced laser desorption/ionization affinity mass spectrometry. This information was used to create a functional mutant of MFE-CP with a significant reduction (range 19.2 to 62.5%, median 38.5%) in reactivity with the sera of 11 patients with post-therapy HACA. The techniques described here are valuable tools for identifying and adapting undesirable immunogenic sites on protein therapeutics.

    更新日期:2019-11-01
  • A model of random mass-matching and its use for automated significance testing in mass spectrometric proteome analysis.
    Proteomics (IF 3.106) Pub Date : 2002-03-29
    Jan Eriksson,David Fenyö

    A rapid and accurate method for testing the significance of protein identities determined by mass spectrometric analysis of protein digests and genome database searching is presented. The method is based on direct computation using a statistical model of the random matching of measured and theoretical proteolytic peptide masses. Protein identification algorithms typically rank the proteins of a genome database according to a score based on the number of matches between the masses obtained by mass spectrometry analysis and the theoretical proteolytic peptide masses of a database protein. The random matching of experimental and theoretical masses can cause false results. A result is significant only if the score characterizing the result deviates significantly from the score expected from a false result. A distribution of the score (number of matches) for random (false) results is computed directly from our model of the random matching, which allows significance testing under any experimental and database search constraints. In order to mimic protein identification data quality in large-scale proteome projects, low-to-high quality proteolytic peptide mass data were generated in silico and subsequently submitted to a database search program designed to include significance testing based on direct computation. This simulation procedure demonstrates the usefulness of direct significance testing for automatically screening for samples that must be subjected to peptide sequence analysis by e.g. tandem mass spectrometry in order to determine the protein identity.

    更新日期:2019-11-01
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