FIG. 6.

Soluble, purified MEKK1 is activated by soluble, purified GCK or TRAF2 in vitro. HA-MEKK1 and FLAG-GCK, GCK(K44M), GCK CTD, and TRAF2 were immunoprecipitated from transfected 293 cells and eluted with the cognate antigen peptide. (A) Silver-stained gels of the purified HA-MEKK1 and FLAG-GCK and TRAF2 constructs. Note that the immunoglobulin G heavy chain [IgG(H)], from the immunoprecipitations, is still present in the eluted protein preparations; however, the free-peptide concentration used in the elutions (100 μM) is far in excess of the IgG concentrations employed in the immunoprecipitation. FLAG-TRAF2 comigrates with the IgG and is not visible in the preparations. (B) Assays for activation of purified MEKK1 by the purified GCK and TRAF2 constructs.