Showing papers by "Jimmy K. Eng published in 2013"
TL;DR: The Comet search engine is introduced, open source, freely available, and based on one of the original sequence database search tools that has been widely used for many years.
Abstract: 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.
1,143 citations
TL;DR: ReACT enables the first view of these interactions inside cells, and the results acquired with this method suggest cross-linking can play a major role in future efforts to map the interactome in cells.
Abstract: Protein interaction topologies are critical determinants of biological function. Large-scale or proteome-wide measurements of protein interaction topologies in cells currently pose an unmet challenge that could dramatically improve understanding of complex biological systems. A primary impediment includes direct protein topology and interaction measurements from living systems since interactions that lack biological significance may be introduced during cell lysis. Furthermore, many biologically relevant protein interactions will likely not survive the lysis/sample preparation and may only be measured with in vivo methods. As a step toward meeting this challenge, a new mass spectrometry method called Real-time Analysis for Cross-linked peptide Technology (ReACT) has been developed that enables assignment of cross-linked peptides “on-the-fly”. Using ReACT, 708 unique cross-linked (<5% FDR) peptide pairs were identified from cross-linked E. coli cells. These data allow assembly of the first protein interact...
135 citations
TL;DR: The results presented here provide new details on the structures of known multi-protein complexes as well as evidence for new protein-protein interactions.
107 citations
TL;DR: XLink-DB serves as a data storage site and visualization tool for cross-linking results and accepts data generated with any cross-linker and stores them in a relational database.
Abstract: As large-scale cross-linking data becomes available, new software tools for data processing and visualization are required to replace manual data analysis. XLink-DB serves as a data storage site and visualization tool for cross-linking results. XLink-DB accepts data generated with any cross-linker and stores them in a relational database. Cross-linked sites are automatically mapped onto PDB structures if available, and results are compared to existing protein interaction databases. A protein interaction network is also automatically generated for the entire data set. The XLink-DB server, including examples, and a help page are available for noncommercial use at http://brucelab.gs.washington.edu/crosslinkdbv1/ . The source code can be viewed and downloaded at https://sourceforge.net/projects/crosslinkdb/?source=directory .
45 citations
TL;DR: The utility of SRM-based targeted proteomics to guide the identification of gene-specific transcriptional regulators at RNA polymerase II transcribed promoters in Saccharomyces cerevisiae is demonstrated.
Abstract: Regulation of gene expression involves the orchestrated interaction of a large number of proteins with transcriptional regulatory elements in the context of chromatin. Our understanding of gene regulation is limited by the lack of a protein measurement technology that can systematically detect and quantify the ensemble of proteins associated with the transcriptional regulatory elements of specific genes. Here, we introduce a set of selected reaction monitoring (SRM) assays for the systematic measurement of 464 proteins with known or suspected roles in transcriptional regulation at RNA polymerase II transcribed promoters in Saccharomyces cerevisiae. Measurement of these proteins in nuclear extracts by SRM permitted the reproducible quantification of 42% of the proteins over a wide range of abundances. By deploying the assay to systematically identify DNA binding transcriptional regulators that interact with the environmentally regulated FLO11 promoter in cell extracts, we identified 15 regulators that bound specifically to distinct regions along ∼600 bp of the regulatory sequence. Importantly, the dataset includes a number of regulators that have been shown to either control FLO11 expression or localize to these regulatory regions in vivo. We further validated the utility of the approach by demonstrating that two of the SRM-identified factors, Mot3 and Azf1, are required for proper FLO11 expression. These results demonstrate the utility of SRM-based targeted proteomics to guide the identification of gene-specific transcriptional regulators.
33 citations
TL;DR: This study identifies a kinase that inhibits Wnt/β-catenin signaling, a pathway critical to melanoma cell viability, and sensitizes melanoma cells to cell death stimulated by WNT3A.
30 citations
01 Jan 2013
TL;DR: These results form the basis for future analyses of alterations in bacterial protein content during growth in various environments, including the cystic cysticrosis airway.
Abstract: proteins expressed during anaerobic growth. Out of the617 proteins identified, 158 were changed in abundance during anaerobic growth compared to during aerobicgrowth, including proteins whose increased expression was expected based on their role in anaerobic metab-olism. These results form the basis for future analyses of alterations in bacterial protein content during growthin various environments, including the cystic fibrosis airway.