Document Type
Article
Version Deposited
Published Version
Publication Date
8-9-2023
Publication Title
STAR Protocols
DOI
10.1016/j.xpro.2023.102489
Abstract
Yeast is an essential model organism for studying protein ubiquitination pathways; however, identifying the direct substrates of E3 in the cell presents a challenge. Here, we present a protocol for using the orthogonal ubiquitin transfer (OUT) cascade to profile the substrate specificity of yeast E3 Rsp5. We describe steps for OUT profiling, proteomics analysis, in vitro and in cell ubiquitination, and stability assay. The protocol can be adapted for identifying and verifying the ubiquitination targets of other E3s in yeast. For complete details on the use and execution of this protocol, please refer to Wang et al.
Recommended Citation
Fang, Shuai; Chen, Geng; Wang, Yiyang; Ganti, Rakhee; Chernova, Tatiana A; Zhou, Li; Jacobs, Savannah E; Duong, Duc; Kiyokawa, Hiroaki; Chernoff, Yury O; Li, Ming; Shcherbik, Natalia; Zhao, Bo; and Yin, Jun, "Profiling and Verifying the Substrates of E3 Ubiquitin Ligase Rsp5 in Yeast Cells" (2023). Rowan-Virtua School of Osteopathic Medicine Departmental Research. 202.
https://rdw.rowan.edu/som_facpub/202
Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial-No Derivative Works 4.0 International License.
Published Citation
Fang S, Chen G, Wang Y, et al. Profiling and verifying the substrates of E3 ubiquitin ligase Rsp5 in yeast cells. STAR Protoc. Published online August 9, 2023. doi:10.1016/j.xpro.2023.102489
Included in
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