Figure 3
From: HIV-1 Vif N-terminal Motif is required for recruitment of Cul5 to Suppress APOBEC3
Select Vif N-terminal mutants have a reduced ability to bind Cul5 in mammalian cells. HA-tagged Vif wild-type and mutant proteins along with CBF-β were overexpressed in HEK 293 T cells. Two days post-transfection, cells were lysed and cleared lysate was mixed with anti-HA matrix affinity beads for 4-8 hrs. Incubated beads were washed several times followed by elution of bound proteins. Select Vif N-terminal mutants (V25A, H27A, M29A, and Y30A) that do not efficiently degrade A3G and A3F have a reduced ability to co-precipitate Cul5; however, CBF-β and Elo B/C can still bind Vif.