Figure 5

Hrd1-mediated K63-linked polyubiquitination of CDV H at position K115. A Schematic of amino-acid sequence of CDV H. Potential ubiquitin lysine residues are displayed in boldface. B CDV H protein with K115R mutation was not degraded by Hrd1. 293 T cells were transfected with the indicated plasmids for 24 h, and then cell lysates were analyzed by Western blot using anti-Flag, anti-Myc and anti-GAPDH antibodies C K115R mutant of CDV H eliminated Hrd1-mediated ubiquitination. 293 T cells were co-transfected with the indicated plasmids for 24 h, and then the cell lysates were subjected to ubiquitination assays and Western blot with anti-Flag, anti-Ha, anti-Myc and anti-GAPDH antibodies. D Schematic of ubiquitin K48 and K63 mutants. E The ubiquitin of CDV H by Hrd1 was K63-linked. 293 T cells were co-transfected with the indicated plasmids for 24 h. The cell lysates were subjected to ubiquitination assays and Western blot with anti-Flag, anti-Ha, anti-Myc and anti-GAPDH antibodies.