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Fig. 1. Lysates were prepared from 293T cells overexpressing (A) Myc-tagged PAK4 alleles or sub domains and incubated with GST, GST-GEF-H1 (787-921) and GST-Maguin-like bound to glutathione beads (1-4 bead designation) or (B) GFP-GEF-H1 and incubated with PAK4 biotinylated peptides (291-355) and (276-324) linked to streptavidin beads. An equivalent amount of beads and excess lysates (500 µg) were used. Eluates of beads and total lysates were analyzed by SDS-PAGE and immunoblotted with Myc or GEF-H1 antiserum. Panel C represents a PAK4 gene schematic outlining the regions of PAK4 used to identify the GEF-H1 interaction domain (GID). (D) Alignment of the GID defined by the minimal binding region of PAK4 with PAK5 and PAK6.





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