FIGURE 1.

pVHL₃₀ migrates as a doublet. (a, b, and d) Biotinylated HIF1α peptides were immobilized on streptavidin‐agarose beads and incubated with in vitro transcribed and translated (IVTT) pVHL in EBC buffer supplemented with protease inhibitors. Streptavidin beads were pulled down (PD), and levels of pVHL were visualized via immmunoblotting (IB). (c) IVTT pVHL was incubated in EBC buffer supplemented with protease inhibitors with or without HIF1α peptides. 3xFLAG‐pVHL was immunoprecipitated (IP) with an anti‐FLAG antibody, and levels of pVHL were visualized via IB. (e) HEK293a cells were transiently transfected HA‐pVHL constructs for 48 hr before being lysed in EBC buffer supplemented with protease inhibitors. Hundred micrograms of total protein was resolved via SDS‐PAGE, and levels of pVHL were visualized via immunoblotting