Fig 6.

Mek1 is affinity purified by Cu-chelated resins. (A) GSH resin, either alone or preincubated and loaded with Cu, was incubated with total protein extracts from wild-type MEFs. The levels of Mek1, GAPDH, and Erk1/2 were assayed from the input proteins, GSH resin affinity-purified proteins, and Cu-charged GSH resin-purified proteins by SDS-PAGE and immunoblotting. (B) Pentadentate-chelated beads complexed with no metal, zinc, or Cu were incubated with lysates from Ctr1^+/+ cells, and Mek1 and the KSR1 scaffold protein were analyzed by immunoblotting as for panel A. (C) Purified recombinant rat Mek1 was added to pentadentate beads that were uncharged (metal free) or charged with zinc (Zn) or Cu, and affinity-purified Mek1 was analyzed by SDS-PAGE and immunoblotting.