The BioGRID | PUBLICATION VIEW

	Mol. Syst. Biol. (2007); 3:116
Ubiquitination screen using protein microarrays for comprehensive identification of Rsp5 substrates in yeast-
Gupta R, Kus B, Fladd C, Wasmuth J, Tonikian R, Sidhu S, Krogan NJ, Parkinson J, Rotin D
Program in Cell Biology, The Hospital for Sick Children, Toronto, Ontario, Canada-
Abstract: Ubiquitin-protein ligases -E3s- are responsible for target recognition and regulate stability, localization or function of their substrates- However, the substrates of most E3 enzymes remain unknown- Here, we describe the development of a novel proteomic in vitro ubiquitination screen using a protein microarray platform that can be utilized for the discovery of substrates for E3 ligases on a global scale- Using the yeast E3 Rsp5 as a test system to identify its substrates on a yeast protein microarray that covers most of the yeast -Saccharomyces cerevisiae- proteome, we identified numerous known and novel ubiquitinated substrates of this E3 ligase- Our enzymatic approach was complemented by a parallel protein microarray protein interaction study- Examination of the substrates identified in the analysis combined with phage display screening allowed exploration of binding mechanisms and substrate specificity of Rsp5- The development of a platform for global discovery of E3 substrates is invaluable for understanding the cellular pathways in which they participate, and could be utilized for the identification of drug targets-
[PUBMED: 17551511]