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proteomics analysis tools

Gregory B Hurst, Keiji G Asano, Charles J Doktycz, Elliot J Consoli, William L Doktycz, Carmen M Foster, Jennifer L Morrell-Falvey, Robert F Standaert, Mitchel J Doktycz
Cell-free protein synthesis (CFPS) has the potential to produce enzymes, therapeutic agents, and other proteins, while circumventing difficulties associated with in vivo heterologous expression. However, the contents of the cell-free extracts used to carry out synthesis are generally not characterized, which hampers progress toward enhancing yield or functional activity of the target protein. We explored the utility of mass spectrometry (MS)-based proteomics for characterizing the bacterial extracts used for transcribing and translating gene sequences into proteins as well as the products of CFPS reactions...
October 17, 2017: Analytical Chemistry
Thierry Bertomeu, Jasmin Coulombe-Huntington, Andrew Chatr-Aryamontri, Karine Bourdages, Etienne Coyaud, Brian Raught, Yu Xia, Mike Tyers
To interrogate genes essential for cell growth, proliferation and survival in human cells, we carried out a genome-wide CRISPR/Cas9 screen in a B-cell lymphoma line using a custom extended knockout (EKO) library of 278,754 sgRNAs that targeted 19,084 RefSeq genes, 20,852 alternatively-spliced exons and 3,872 hypothetical genes. A new statistical analysis tool called RANKS identified 2,280 essential genes, 234 of which were unique. Individual essential genes were validated experimentally and linked to ribosome biogenesis and stress responses...
October 16, 2017: Molecular and Cellular Biology
Svetlana E Novikova, Olga V Tikhonova, Leonid K Kurbatov, Tatiana E Farafonova, Igor V Vakhrushev, Victor G Zgoda
Targeted mass spectrometry represents a powerful tool for investigation of biological processes. The convenient approach of selected reaction monitoring using stable isotope-labeled peptide standard (SIS) is widely applied for protein quantification. Along with this method, high-resolution parallel reaction monitoring has been increasingly used for protein targeted analysis. Here we applied two targeted approaches (selected reaction monitoring with SIS and label-free parallel reaction monitoring) to investigate expression of 11 proteins during all-trans retinoic acid-induced differentiation of HL-60 cells...
August 2017: European Journal of Mass Spectrometry
Tian Qiu, Rahul S Kathayat, Yang Cao, Michael William Beck, Bryan C Dickinson
S-palmitoylation is an abundant lipid post-translational modification (PTM) that is dynamically installed on and removed from target proteins to regulate their activity and cellular localization. A dearth of tools to study the activities and regulation of protein S-depalmitoylases - thioesterase "erasers" of protein cysteine S-palmitoylation - has contributed to an incomplete understanding of the role of dynamic S-palmitoylation in regulating proteome lipidation. Recently, we developed "depalmitoylation probes" (DPPs), small molecule probes that become fluorescent upon S-depalmitoylase enzymatic activity...
October 12, 2017: Biochemistry
Azmeraw T Amare, Klaus Oliver Schubert, Bernhard T Baune
Personalized medicine (personalized psychiatry in a specific setting) is a new model towards individualized care, in which knowledge from genomics and other omic pillars (microbiome, epigenomes, proteome, and metabolome) will be combined with clinical data to guide efforts to new drug development and targeted prescription of the existing treatment options. In this review, we summarize pharmacogenomic studies in mood disorders that may lay the foundation towards personalized psychiatry. In addition, we have discussed the possible strategies to integrate data from omic pillars as a future path to personalized psychiatry...
September 2017: EPMA Journal
Jarlath E Nally, Simone Schuller
Lungs perform an essential physiological function, mediated by a complex series of events that involve the coordination of multiple cell types to support not only gaseous exchange, but homeostasis and protection from infection. Guinea pigs are an important animal disease model for a number of infectious and noninfectious pulmonary conditions and the availability of a complete genome facilitates comprehensive analysis of tissues using the tools of proteomics. Here, we describe the application of 2-D Difference Gel Electrophoresis (DIGE) to compare, quantify, and identify differential protein expression of proteins in lung tissue from guinea pigs with leptospiral pulmonary hemorrhage syndrome (LPHS) compared to noninfected controls...
2018: Methods in Molecular Biology
Kay Ohlendieck
The skeletal muscle proteome consists of a large number of diverse protein species with a broad and dynamic concentration range. Since mature skeletal muscles are characterized by a specific combination of contractile cells with differing physiological and biochemical properties, it is essential to determine specific differences in the protein composition of fast, slow, and hybrid fibers. Fluorescence two-dimensional gel electrophoresis (DIGE) is a powerful comparative tool to analyze fiber type-specific differences between fast and slow muscles...
2018: Methods in Molecular Biology
Xiaoyu Ji, Xiaoqiang Liu, Yuanxia Peng, Ruoting Zhan, Hui Xu, Xijin Ge
Emodin has a strong antibacterial activity, including methicillin-resistant Staphylococcus aureus (MRSA). However, the mechanism by which emodin induces growth inhibition against MRSA remains unclear. In this study, the isobaric tags for relative and absolute quantitation (iTRAQ) proteomics approach was used to investigate the modes of action of emodin on a MRSA isolate and methicillin-sensitive S. aureus ATCC29213(MSSA). Proteomic analysis showed that expression levels of 145 and 122 proteins were changed significantly in MRSA and MSSA, respectively, after emodin treatment...
October 7, 2017: Biochemical and Biophysical Research Communications
Nicolas F Fernandez, Gregory W Gundersen, Adeeb Rahman, Mark L Grimes, Klarisa Rikova, Peter Hornbeck, Avi Ma'ayan
Most tools developed to visualize hierarchically clustered heatmaps generate static images. Clustergrammer is a web-based visualization tool with interactive features such as: zooming, panning, filtering, reordering, sharing, performing enrichment analysis, and providing dynamic gene annotations. Clustergrammer can be used to generate shareable interactive visualizations by uploading a data table to a web-site, or by embedding Clustergrammer in Jupyter Notebooks. The Clustergrammer core libraries can also be used as a toolkit by developers to generate visualizations within their own applications...
October 10, 2017: Scientific Data
Ahmad F Karim, Obondo J Sande, Sara E Tomeckho, Xuedong Ding, Ming Li, Sean Maxwell, Rob M Ewing, Clifford V Harding, Roxana E Rojas, Mark R Chance, W Henry Boom
Mycobacterium tuberculosis (Mtb) cell wall glycolipid Mannose-capped Lipoarabinomannan (ManLAM) inhibits CD4(+) T cell activation by inhibiting proximal T cell receptor (TCR) signaling when activated by anti-CD3. To understand the impact of ManLAM on CD4(+) T cell function when both the TCR-CD3 complex and major co-stimulator CD28 are engaged, we performed label-free quantitative mass spectrometry (MS) and network analysis. Mixed effect model analysis of peptide intensity identified 149 unique peptides representing 131 proteins that were differentially regulated by ManLAM in anti-CD3 and anti-CD28 activated CD4(+) T cells...
October 10, 2017: Proteomics
Kristina Poljak, Nathalie Selevsek, Elsy Ngwa, Jonas Grossmann, Marie Estelle Losfeld, Markus Aebi
Asparagine-linked glycosylation is a common posttranslational protein modification regulating the structure, stability and function of many proteins. The N-linked glycosylation machinery involves enzymes responsible for the assembly of the lipid-linked oligosaccharide (LLO), which is then transferred to the asparagine residues on the polypeptides by the enzyme oligosaccharyltransferase (OST).  A major goal in the study of protein glycosylation is to establish quantitative methods for the analysis of site-specific extent of glycosylation...
October 9, 2017: Molecular & Cellular Proteomics: MCP
Paolo Soffientini, Angela Bachi
Proteomics is nowadays a standard tool in life sciences for the analysis of protein abundance, modifications and interactions but has so far failed to enter the clinic for routine applications. New generation mass spectrometers and chromatographic systems are able to cover approximately an entire cell proteome in one run but sample preparation, in terms of time and sample recovery, is still a critical step. Here we describe a modification of the in-gel digestion method, called STAGE-diging, that reduces sample handling, decreases the analysis time and improves protein identification and quantification...
2017: Methods in Molecular Biology
Alejandro Brenes, Vackar Afzal, Robert Kent, Angus I Lamond
Driven by improvements in speed and resolution of mass spectrometers (MS), the field of proteomics, which involves the large-scale detection and analysis of proteins in cells, tissues and organisms, continues to expand in scale and complexity. There is a resulting growth in datasets of both raw MS files and processed peptide and protein identifications. MS-based proteomics technology is also used increasingly to measure additional protein properties affecting cellular function and disease mechanisms, including post-translational modifications, protein-protein interactions, subcellular and tissue distributions...
September 7, 2017: Nucleic Acids Research
Ting Cao, Lei Zhang, Ying Zhang, Guoquan Yan, Caiyun Fang, Huimin Bao, Haojie Lu
Proteome-wide quantitative analysis of protein ubiquitination is important to gain insight into its various cellular functions. However, it is still challenging to monitor how ubiquitination at each individual lysine residue is independently regulated, especially the whereabouts of peptides containing more than one ubiquitination site. In recent years, isobaric peptide termini labeling has been considered a promising strategy in quantitative proteomics, benefiting from its high accuracy by quantifying with a series of b, y fragment ion pairs...
October 18, 2017: Analytical Chemistry
Achim Treumann, Pawel Palmowski, Wing Chiu Tong, Julie Taggart, Nick Morrice, G Nicholas Europe-Finner, Michael J Taggart
Muscle tissue poses a particular challenge to proteomic analysis due to a very wide range of protein abundances arising from the dominant expression of myofilament-related proteins. We address this issue by describing proteomic analysis with liquid chromatography-mass spectrometry (LC-MS) and sequential window acquisition of all theoretical mass spectra (SWATH), of guinea pig cardiac tissue prepared in two homogenization buffers: (1) An SDS-based buffer designed to extract "all" tissue proteins and (2) a long-established EDTA-containing buffer thought to preferentially extract non-myofibril-related proteins...
October 4, 2017: Methods in Molecular Biology
J F Chignell, S Park, C M R Lacerda, S K De Long, K F Reardon
Interactions among members of microbial consortia drive the complex dynamics in soil, gut, and biotechnology microbiomes. Proteomic analysis of defined co-cultures of well-characterized species provides valuable information about microbial interactions. We used a label-free approach to quantify the responses to co-culture of two model bacterial species relevant to soil and rhizosphere ecology, Bacillus atrophaeus and Pseudomonas putida. Experiments determined the ratio of species in co-culture that would result in the greatest number of high-confidence protein identifications for both species...
October 3, 2017: Microbial Ecology
Suowen Xu
In the post-genomic, big data era, our understanding of vascular diseases has been deepened by multiple state-of-the-art "-omics" approaches, including genomics, epigenomics, transcriptomics, proteomics, lipidomics and metabolomics. Genome-wide transcriptomic profiling, such as gene microarray and RNA-sequencing, emerges as powerful research tools in systems medicine and revolutionizes transcriptomic analysis of the pathological mechanisms and therapeutics of vascular diseases. In this article, I will highlight the workflow of transcriptomic profiling, outline basic bioinformatics analysis, and summarize recent gene profiling studies performed in vascular cells as well as in human and mice diseased samples...
2017: Frontiers in Pharmacology
Ngoc Q Vuong, Dalibor Breznan, Patrick Goegan, Julie S O'Brien, Andrew Williams, Subramanian Karthikeyan, Premkumari Kumarathasan, Renaud Vincent
BACKGROUND: Toxicity of airborne particulate matter (PM) is difficult to assess because PM composition is complex and variable due to source contribution and atmospheric transformation. In this study, we used an in vitro toxicoproteomic approach to identify the toxicity mechanisms associated with different subfractions of Ottawa urban dust (EHC-93). METHODS: A549 human lung epithelial cells were exposed to 0, 60, 140 and 200 μg/cm(2) doses of EHC-93 (total), its insoluble and soluble fractions for 24 h...
October 2, 2017: Particle and Fibre Toxicology
Elena Papaleo, Irina Gromova, Pavel Gromov
Tumor-associated proteins released by cancer cells and by tumor stroma cells, referred as "cancer secretome", represent a valuable resource for discovery of potential cancer biomarkers. The last decade was marked by a great increase in number of studies focused on various aspects of cancer secretome including, composition and identification of components externalized by malignant cells and by the components of tumor microenvironment. Areas covered: Here, we provide an overview of achievements in the proteomic analysis of the cancer secretome, elicited through the tumor-associated interstitial fluid recovered from malignant tissues ex vivo or the protein component of conditioned media obtained from cultured cancer cells in vitro...
October 2, 2017: Expert Review of Proteomics
Pierre H Boyer, Nathalie Boulanger, Amira Nebbak, Elodie Collin, Benoit Jaulhac, Lionel Almeras
Matrix Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) has been demonstrated to be useful for tick identification at the species level. More recently, this tool has been successfully applied for the detection of bacterial pathogens directly in tick vectors. The present work has assessed the detection of Borrelia burgdorferi sensu lato in Ixodes ricinus tick vector by MALDI-TOF MS. To this aim, experimental infection model of I. ricinus ticks by B. afzelii was carried out and specimens collected in the field were also included in the study...
2017: PloS One
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