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Realised by ALMS™
developer of the AIDS-HIV Reference project
Abstract No.: TuOr-18
Speaker: James E. Bruce
Session: Peptidomics and Proteomics
Presentation date: Tue, Aug 29, 2006
Presentation time: 16:30 – 16:50

New Technology for Protein Interaction Network and Topology Analysis

James E. Bruce1, Xiaoting Tang1, Wei Yi1, Saiful Chowdhury1, Gerhard Munske1, Gordon Anderson2, Nikola Tolic2

1 Washington State University, Pullman, United States
2 EMSL, Richland, United States

Correspondence address: James E. Bruce, Washington State University, Chemistry, College Blvd, Pullman, 99164-4630 United States.

Keywords: Cross-linking; Dissociation; Fourier Transform ICR; Labeling.

Novel aspect: Novel method for protein interaction network identification.

 

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 an unmet challenge for today's technology. Improved capabilities to monitor and visualize these networks will have a major impact in the current understanding of many diseases, including cancer, diabetes, cardiovascular disease and virtually all areas of human health. We are developing new 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 covalently cross-links proteins in cells and facilitates affinity capture of labeled proteins. The novel, key feature of the PIR system is based on incorporation releasable reporter ions that are 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 pinpoint 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.
This presentation will highlight our recent efforts to develop the PIR strategy for protein interaction and topology analysis with intact cells. We will present our protein identification efforts with the bacterial system, Shewanella oneidensis MR-1, that resulted in the identification of more than 375 proteins that were labeled and identified with our PIRs. We will also demonstrate observed differences in proteins that were identified with variation of PIR structures. The cleavable features of the PIR structures were exploited to allow accurate mass and tandem MS-based identification of peptides and sites of incorporation. This information allows identification of solvent exposed protein surfaces, as well as interactions among proteins in cells.