Proteomics research routinely involves identifying peptides and proteins via MS/MS sequence database search. Thus the database search engine is an integral tool in many proteomics research groups. ...Here, we introduce the Comet search engine to the existing landscape of commercial and open‐source database search tools. Comet is open source, freely available, and based on one of the original sequence database search tools that has been widely used for many years.
Multiplexed quantitative analyses of complex proteomes enable deep biological insight. While a multitude of workflows have been developed for multiplexed analyses, the most quantitatively accurate ...method (SPS-MS3) suffers from long acquisition duty cycles. We built a new, real-time database search (RTS) platform, Orbiter, to combat the SPS-MS3 method’s longer duty cycles. RTS with Orbiter eliminates SPS-MS3 scans if no peptide matches to a given spectrum. With Orbiter’s online proteomic analytical pipeline, which includes RTS and false discovery rate analysis, it was possible to process a single spectrum database search in less than 10 ms. The result is a fast, functional means to identify peptide spectral matches using Comet, filter these matches, and more efficiently quantify proteins of interest. Importantly, the use of Comet for peptide spectral matching allowed for a fully featured search, including analysis of post-translational modifications, with well-known and extensively validated scoring. These data could then be used to trigger subsequent scans in an adaptive and flexible manner. In this work we tested the utility of this adaptive data acquisition platform to improve the efficiency and accuracy of multiplexed quantitative experiments. We found that RTS enabled a 2-fold increase in mass spectrometric data acquisition efficiency. Orbiter’s RTS quantified more than 8000 proteins across 10 proteomes in half the time of an SPS-MS3 analysis (18 h for RTS, 36 h for SPS-MS3).
Understanding metabolic dysregulation in different disease settings is vital for the safe and effective incorporation of metabolism-targeted therapeutics in the clinic. Here, using transcriptomic ...data for 10,704 tumor and normal samples from The Cancer Genome Atlas, across 26 disease sites, we present a novel bioinformatics pipeline that distinguishes tumor from normal tissues, based on differential gene expression for 114 metabolic pathways. We confirm pathway dysregulation in separate patient populations, demonstrating the robustness of our approach. Bootstrapping simulations were then applied to assess the biological significance of these alterations. We provide distinct examples of the types of analysis that can be accomplished with this tool to understand cancer specific metabolic dysregulation, highlighting novel pathways of interest, and patterns of metabolic flux, in both common and rare disease sites. Further, we show that Master Metabolic Transcriptional Regulators explain why metabolic differences exist, can segregate patient populations, and predict responders to different metabolism-targeted therapeutics.
Hsp90 belongs to a family of some of the most highly expressed heat shock proteins that function as molecular chaperones to protect the proteome not only from the heat shock but also from other ...misfolding events. As many client proteins of Hsp90 are involved in oncogenesis, this chaperone has been the focus of intense research efforts. Yet, we lack structural information for how Hsp90 interacts with co-chaperones and client proteins. Here, we developed a mass-spectrometry-based approach that allowed quantitative measurements of in vitro and in vivo effects of small-molecule inhibitors on Hsp90 conformation, and interaction with co-chaperones and client proteins. From this analysis, we were able to derive structural models for how Hsp90 engages its interaction partners in vivo, and how different drugs affect these structures. In addition, the methodology described here offers a new approach to probe the effects of virtually any inhibitor treatment on the proteome level.
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•Hsp90 conformational changes upon inhibitor treatment in cells revealed•Hsp90-isoform-specific responses to inhibitor treatment visualized•Quantitative Hsp90 interactome network including 3,323 crosslinked peptides•Structural information provided for Hsp90 co-chaperone and client protein complexes
Hsp90 functions to maintain cellular homeostasis. Chavez et al. identified in vivo changes to Hsp90 conformations and interactions upon cellular treatment with Hsp90 inhibitors using quantitative crosslinking with mass spectrometry. Conformational changes were found to be drug and isoform specific.
We demonstrate improved operation of exchange-coupled semiconductor quantum dots by substantially reducing the sensitivity of exchange operations to charge noise. The method involves biasing a double ...dot symmetrically between the charge-state anticrossings, where the derivative of the exchange energy with respect to gate voltages is minimized. Exchange remains highly tunable by adjusting the tunnel coupling. We find that this method reduces the dephasing effect of charge noise by more than a factor of 5 in comparison to operation near a charge-state anticrossing, increasing the number of observable exchange oscillations in our qubit by a similar factor. Performance also improves with exchange rate, favoring fast quantum operations.
Protein identification by tandem mass spectrometry sequence database searching is a standard practice in many proteomics laboratories. The de facto standard for the representation of sequence ...databases used as input to sequence database search tools is the FASTA format. The Human Proteome Organization's Proteomics Standards Initiative has developed an extension to the FASTA format termed the proteomics standards initiative extended FASTA format or PSI extended FASTA format (PEFF) where additional information such as structural annotations are encoded in the protein description lines. Comet has been extended to automatically analyze the post translational modifications and amino acid substitutions encoded in PEFF databases. Comet's PEFF implementation and example analysis results searching a HEK293 dataset against the neXtProt PEFF database are presented.
The combination of tandem mass spectrometry and sequence database searching is the method of choice for the identification of peptides and the mapping of proteomes. Over the last several years, the ...volume of data generated in proteomic studies has increased dramatically, which challenges the computational approaches previously developed for these data. Furthermore, a multitude of search engines have been developed that identify different, overlapping subsets of the sample peptides from a particular set of tandem mass spectrometry spectra. We present iProphet, the new addition to the widely used open-source suite of proteomic data analysis tools Trans-Proteomics Pipeline. Applied in tandem with PeptideProphet, it provides more accurate representation of the multilevel nature of shotgun proteomic data. iProphet combines the evidence from multiple identifications of the same peptide sequences across different spectra, experiments, precursor ion charge states, and modified states. It also allows accurate and effective integration of the results from multiple database search engines applied to the same data. The use of iProphet in the Trans-Proteomics Pipeline increases the number of correctly identified peptides at a constant false discovery rate as compared with both PeptideProphet and another state-of-the-art tool Percolator. As the main outcome, iProphet permits the calculation of accurate posterior probabilities and false discovery rate estimates at the level of sequence identical peptide identifications, which in turn leads to more accurate probability estimates at the protein level. Fully integrated with the Trans-Proteomics Pipeline, it supports all commonly used MS instruments, search engines, and computer platforms. The performance of iProphet is demonstrated on two publicly available data sets: data from a human whole cell lysate proteome profiling experiment representative of typical proteomic data sets, and from a set of Streptococcus pyogenes experiments more representative of organism-specific composite data sets.
Service innovation is focused on customer value creation. At its core, customer centric service innovation is technology-enabled, human-centered, and process-oriented. To profit from such innovation, ...firms need an
integrated
cross-disciplinary, holistic method to design
and
commercialize service innovation. From diverse but interrelated strands of theories from service science, strategic management, organization science and information systems literatures, this article develops a new integrated design method, known as
i
SIM (
integrated
Service Innovation Method), for simultaneous service innovation
and
business model design for sustained customer value co-creation with the firm. Following design science research method, the article theoretically defines and integrates
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SIM’s seven constitutive design process-elements: service strategy, customer type / value proposition, service concept, service system, customer experience, service architecture and monetization into a coherent and end-to-end aligned integrated design method. It explains how
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SIM would be holistically and iteratively practiced by practitioners, and conceptually exemplifies its utility via telco and Amazon case studies using secondary data. Perspectives on
i
SIM from selected practitioners are discussed which confirm
i
SIM’s potential utility for their business. Managerial implications of implementing the
i
SIM and potential areas for further research are also discussed.
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