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. 2010 Oct 27;5(10):e13695. doi: 10.1371/journal.pone.0013695

Figure 3. 2UIM and 3UIM ataxin-3 display similar DUB activity against defined ubiquitin chains in vitro.

Figure 3

(A–C) Recombinant GST-ATXN3(Q22) (3UIM or 2UIM) can cleave K48-linked hexaubiquitin (A), K63-linked tetraubiquitin (B), and mixed linkage K48-K63-K48 tetraubiquitin (C) chains. Results with catalytically inactive GST-ataxin-3 (C14A mutant) are also shown. (D–E) Recombinant 2UIM and 3UIM GST-ATXN3(Q22) cleave Ub-AMC at a similar rate. (D) Ub-AMC reaction curves. Both 3UIM and 2UIM ataxin-3 area able to cleave Ub-AMC, while reactions with either an unrelated control protein (the non-DUB F-box protein FBXO2) or buffer only show no cleavage. Error bars show standard deviations. (E) There is no significant difference between the initial reaction velocity of 2UIM and 3UIM ataxin-3(Q22) (p>0.4 by a 2 tailed heteroscedastic Student's t-test).