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United States Department of Agriculture

Agricultural Research Service

Research Project: GENOMICS AND PROTEOMICS APPROACHES TO BROADENING RESISTANCE OF SOYBEAN TO PESTS AND PATHOGENS Title: Affinity purification and mass spectrometry: an attractive choice to investigate protein-protein interactions in plant immunity

Authors
item Chen, Ruiqiang -
item Chang, Caren -
item Tucker, Mark
item Cooper, Bret

Submitted to: Current Protemics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: April 16, 2010
Publication Date: June 24, 2010
Repository URL: http://handle.nal.usda.gov/10113/56694
Citation: Chen, R., Chang, C., Tucker, M.L., Cooper, B. 2010. Affinity purification and mass spectrometry: an attractive choice to investigate protein-protein interactions in plant immunity. Current Protemics. 7:258-264.

Interpretive Summary: Early research on proteins and their function clearly indicated that proteins must interact to perform their normal function in biological systems. However, it’s only in recent years that technologies have been developed to identify one-on-one protein interactions and the interactions of multiple proteins in a protein complex. One approach that is particularly useful for identifying all the protein partners in a protein complex is a technique called AP-MS/MS that includes affinity purification of the protein complex followed by identification of the proteins in the complex using a device called a tandem mass spectrometer. High throughput identification of proteins by mass spectrometry has only recently become possible due to a large increase in genetic (DNA) sequence data. To take advantage of these new capabilities, protocols for the purification of interacting protein complexes from plant tissue have been improved on and software developed to analyze and interpret the MS/MS data in regard to predictions made based on the DNA sequence data. The recent advances in AP-MS/MS technologies described here will be useful for scientists in academia, government and industry.

Technical Abstract: Affinity purification of protein complexes from biological tissues, followed by liquid chromatography- tandem mass spectrometry (AP-MS/MS), has ballooned in recent years due to sizeable increases in nucleic acid sequence data essential for interpreting mass spectra, improvements in affinity purification protocols and easier-to-use mass spectrometers. Plant biologists have noticed these trends and are now using AP-MS/MS to identify plausible protein-protein interactions crucial to plant defense systems. As a result, knowledge of protein interactions in plants has grown. For example, new partners have been found to interact with RIN4 and RPS2, two plasma membrane-bound proteins critical for defense responses in Arabidopsis thaliana. Moreover, a nuclear protein complex in A. thaliana that includes the defense signaling protein MOS4 has been purified and many of the protein partners that were identified shown to be conserved in a protein complex found in yeast and humans. In another example, several proteins were identified that interact with a defense signaling GTPase, Rac1, in rice. Clearly, AP-MS/MS is an important technique that will continue to provide novel insight into protein interaction networks in plants. These recent advances and the different techniques of AP-MS/MS described here will be useful to plant pathologists in the study of plant-pest interactions.

Last Modified: 10/21/2014
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