Figure 5.

RNF126-Mediated Ubiquitination Is Specific to MLPs

(A) The Bag6-Luc complex was incubated with E1, E2, ATP, Flag-ubiquitin, HEK293T lysate, and recombinant RNF126F at final concentrations of 2.2, 4.4, 8.8, and 22 μM. The products were analyzed after ubiquitin pull-downs by immunoblotting for Luc. Bottom panel shows Coomassie-stained gel of added proteins. GST was used as a control.

(B) PrP or ΔΔPrP was translated in the presence or absence of 4.4 μM RNF126F. The products were visualized directly (bottom) after ubiquitin pull-downs (Ub-PD). Relative ubiquitination in the presence of RNF126F (mean ± SD, n = 3) is indicated below each panel.

(C) Sec61β or Sec61β-3R was translated and analyzed as in (B).

(D) Ub-GFP was translated and analyzed as in (B).

See also Figure S5.