Protein interaction networks are key determinants of protein function in biological systems. However, global or large-scale analysis of protein interaction networks within native living systems is a challenge that is unmet by today's technology. Improved capabilities to monitor and visualize these networks will have a major impact in the understanding of many diseases including cancer, diabetes, cardiovascular disease and virtually all areas of human health. This project aims to develop enabling technology for improved characterization of protein interaction networks and protein topological features in native cellular environments. This advance is based on the new concepts of a Protein Interaction Reporter (PIR) system that functions by cross-linking proteins in cells and facilitates affinity capture of labeled proteins. The novel, key feature of the PIR system is based on incorporation of a releasable reporter ion that is detected during mass spectrometry analysis. The reporter ions are mass-encoded to facilitate information retrieval from complex cross-linking reaction mixtures that will simultaneously include a wide variety of PIR structures, lengths, physical properties and reactive functionalities. The reporter ions allow product differentiation and protein identification, establish connectivity among cross-linked proteins, and pinpoints sites of protein interactions and exposed protein residues that are present within the cellular environment. The development of functional PIR technology that can enable improved network and topological features to be visualized within cells will significantly improve the ability to understand critical aspects of global protein function relevant to human health.

National Institute of Health (NIH)
National Center for Research Resources (NCRR)
Research Project (R01)
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Enabling Bioanalytical and Biophysical Technologies Study Section (EBT)
Program Officer
Sheeley, Douglas
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University of Washington
Schools of Medicine
United States
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Chavez, Juan D; Schweppe, Devin K; Eng, Jimmy K et al. (2016) In Vivo Conformational Dynamics of Hsp90 and Its Interactors. Cell Chem Biol 23:716-26
Chavez, Juan D; Eng, Jimmy K; Schweppe, Devin K et al. (2016) A General Method for Targeted Quantitative Cross-Linking Mass Spectrometry. PLoS One 11:e0167547
Chavez, Juan D; Schweppe, Devin K; Eng, Jimmy K et al. (2015) Quantitative interactome analysis reveals a chemoresistant edgotype. Nat Commun 6:7928
Chowdhury, Saiful M; Munske, Gerhard R; Yang, Jonathon et al. (2014) Solid-phase N-terminal peptide enrichment study by optimizing trypsin proteolysis on homoarginine-modified proteins by mass spectrometry. Rapid Commun Mass Spectrom 28:635-44
Zheng, Chunxiang; Weisbrod, Chad R; Chavez, Juan D et al. (2013) XLink-DB: Database and Software Tools for Storing and Visualizing Protein Interaction Topology Data. J Proteome Res :
Weisbrod, Chad R; Hoopmann, Michael R; Senko, Michael W et al. (2013) Performance evaluation of a dual linear ion trap-Fourier transform ion cyclotron resonance mass spectrometer for proteomics research. J Proteomics 88:109-19
Tamborindeguy, Cecilia; Bereman, Michael S; DeBlasio, Stacy et al. (2013) Genomic and proteomic analysis of Schizaphis graminum reveals cyclophilin proteins are involved in the transmission of cereal yellow dwarf virus. PLoS One 8:e71620
Chavez, Juan D; Weisbrod, Chad R; Zheng, Chunxiang et al. (2013) Protein interactions, post-translational modifications and topologies in human cells. Mol Cell Proteomics 12:1451-67
Chavez, Juan D; Cilia, Michelle; Weisbrod, Chad R et al. (2012) Cross-linking measurements of the Potato leafroll virus reveal protein interaction topologies required for virion stability, aphid transmission, and virus-plant interactions. J Proteome Res 11:2968-81
Weisbrod, Chad R; Eng, Jimmy K; Hoopmann, Michael R et al. (2012) Accurate peptide fragment mass analysis: multiplexed peptide identification and quantification. J Proteome Res 11:1621-32

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