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. 2019 Dec 1;33(23-24):1702–1717. doi: 10.1101/gad.332395.119

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© 2019 Wu et al.; Published by Cold Spring Harbor Laboratory Press

This article is distributed exclusively by Cold Spring Harbor Laboratory Press for the first six months after the full-issue publication date (see http://genesdev.cshlp.org/site/misc/terms.xhtml). After six months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.

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Figure 1. — Identification of Cezanne and Cezanne2 UBA domain as a K63-linked ubiquitin binding motif. (A) A protein array screen identifies the K63-ubiquitin binding motif. Cy3-streptavidin-labeled biotin-K63- and K48- tetra-ubiquitin were used to probe the UBD arrays. The positions of selective K63-linked ubiquitin chain binding protein (domain) are marked: 1-ASCC2 (CUE); 2-Cezanne (UBA); 3-Cezanne2 (UBA); 4-TDRD3 (UBA); 5-RAP80 (UIMs); 6-TOM1L2 (VHS); and 7-RNF168 (MIU2). (B) In vitro pull-down assay using purified recombinant GST-Cezanne UBA, Cezanne2 UBA, or Rap80 UIMs incubated with K63-, K48-, or K11-tetra-ubiquitin chains. Anti-ubiquitin antibody was used for the western blot. (C) ITC analysis of Cezanne, Cezanne2 UBA, or Rap80 UIMs binding to K63-di-Ub.