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. 2009 May 15;284(20):13301–13305. doi: 10.1074/jbc.C900009200

FIGURE 2.

FIGURE 2.

CCM2 is not a Smurf1 substrate, nor does it impact Smurf1 catalytic activity. A, cells expressing HA-CCM2, HA-MEKK2, Myc-RhoA, FLAG-Smurf1 C699A, or FLAG-Smurf1 WT were lysed, and steady-state levels of HA-CCM2, HA-MEKK2, or Myc-RhoA were determined by Western blot. B, cells expressing FLAG-Smurf1 C699A, FLAG-Smurf1 WT, or HA-CCM2 were lysed, and FLAG-Smurf1 proteins were immunopurified (IP). The immunopurified FLAG-Smurf1 or FLAG-Smurf1 C699A was subsequently used for an in vitro ubiquitination assay. C, cells expressing FLAG-Smurf1 C699A, FLAG-Smurf1 WT, or FLAG-Smurf1 YY-AA were lysed, and immunopurified FLAG-Smurf1 was used for in vitro ubiquitination.