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. 2015 Jul 3;16(9):1131–1144. doi: 10.15252/embr.201540298

Figure 2.

Figure 2

In vitro characterization of ubiquitin chain assembly by S65 ubiquitin mutants

Anti-ubiquitin immunoblots of in vitro reactions of ubiquitylation machinery with S65E, S65A, or WT recombinant ubiquitin.
  1. Loading of recombinant ubiquitin mutants onto an Uba1 E1 enzyme.
  2. Generation of unanchored ubiquitin chains using the Ubc1 E2 enzyme.
  3. Generation of unanchored ubiquitin chains using the Ubc13/Mms2 E2 enzyme complex.
  4. Rsp5 HECT domain auto-ubiquitylation using the Ubc4 E2 enzyme.