Figure 6 | Experimental & Molecular Medicine

Figure 6

From: Ubiquitylation of Fe65 adaptor protein by neuronal precursor cell expressed developmentally down regulated 4-2 (Nedd4-2) via the WW domain interaction with Fe65

Figure 6

Ubiquitylation of Fe65 dependent on Nedd4-2 concentration. (A) The immunopurified Fe65 protein with EGFP Ab from HEK 293 was immunoblotted with anti-Fe65 anti-ubiquitin monoclonal antibody. The same cell lysate was immunoblotted with anti-Ha to monitor the Ha Nedd4-2 protein amount. DNA amount of Ha Nedd4-2 (top lane) and that of EGFP Fe65 (wt) were indicated. (B) The immunopurified Nedd4-2 was used as E3 ligase for the in vitro ubiquitinizion assay. The purified GST Fe65 (wt) was ubiqutinized (the right lane), whereas GST-Fe65 W26R mutant was less ubiquitylated, suggesting that K26 residue of Fe65 is one of its major ubiquitylation sites by Nedd4-2. The relative optical density by image analysis with the Fuji Image Quant software is indicated.

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