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Fig. 5 | Parasites & Vectors

Fig. 5

From: An OTU deubiquitinating enzyme in Eimeria tenella interacts with Eimeria tenella virus RDRP

Fig. 5

Et-OTU DUB activity was enhanced by Etv-RDRP/Et-OTU complexes in vitro. In vitro deubiquitination assays were performed using Etv-RDRP in combination with Et-OTU and the Et-OTU protein purified from Sf9 as previously described. K48- and K6-linked di-Ub chains were individually used as substrates at a final concentration of 5 μM. The final concentrations of Et-OTU and Etv-RDRP in combination with Et-OTU in the reaction were both 5 μM. a Etv-RDRP in combination with Et-OTU and the Et-OTU protein was purified using glutathione-Sepharose 4B and individually analysed by Western blot. b In total, 10 μl of both purified Et-OTU DUB and purified RDRP/OTU complexes were separately incubated with 10 μl of K48-linked di-Ub chains for 30 min and resolved by Western blotting. The hydrolytic activity of Et-OTU DUB towards K48-linked di-Ub chains was analysed using ImageJ software. c In total, 10 μl of both purified Et-OTU DUB and purified RDRP/OTU complexes were separately incubated with 10 μl of K6-linked di-Ub chains for 30 min and resolved by Western blotting. The hydrolytic activity of Et-OTU DUB towards K6-linked di-Ub chains was analysed using ImageJ software

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