Reference: Wykoff DD and O'Shea EK (2005) Identification of sumoylated proteins by systematic immunoprecipitation of the budding yeast proteome. Mol Cell Proteomics 4(1):73-83

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Abstract

The identification of post-translational modifications to proteins is critical for understanding many important aspects of biology. Utilizing a collection of epitope-tagged yeast strains, we developed a novel approach to determine which proteins are modified by the small ubiquitin related modifier, SUMO. We crossed traits useful for the detection of SUMO conjugation into 4246 TAP-tagged strains and successfully immunoprecipitated and screened 2893 of these proteins for association with SUMO (approximately 70% of the expressed proteome detectable by immunoblot analysis). We found 82 proteins associated with SUMO, including many of low abundance. Because our screen was performed under non-denaturing conditions, we were able to identify multiple members of four complexes that were associated with SUMO: the RSC chromatin remodeling complex, the mediator complex, the TFIID complex, and the septin complex. In addition, we describe 5 new direct conjugates of SUMO, and we mutated SUMO conjugation sites in four proteins. This is the first attempt to immunoprecipitate a large fraction of the proteome of a eukaryote, and it demonstrates the utility of this method to identify post-translational modifications in the yeast proteome.

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Journal Article
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Wykoff DD, O'Shea EK
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