FIGURE 4.

The IQ region of IQGAP1 is both necessary and sufficient for HER2 binding. A, shown is a schematic representation of IQGAP1 constructs depicting full-length IQGAP1, truncated IQGAP1 fragments, and a deletion mutant. The specific amino acids absent from each construct are indicated. FL, full-length IQGAP1 (amino acids 2–1657); N, N-terminal half of IQGAP1 (amino acids 2–863); IQ, IQ region of IQGAP1 (amino acids 717–916); C, C-terminal half of IQGAP1 (amino acids 864–1657); ΔIQ, full length IQGAP1 from which the IQ region has been deleted. B, [³⁵S]methionine-labeled full length IQGAP1 (FL), IQGAP1-N (N), IQGAP1-IQ (IQ), IQGAP1-C (C), and IQGAP1ΔIQ (ΔIQ) were incubated with equal amounts of GST-HER2 (HER2, top right panel), GST-Cdc42V12 (Cdc42, bottom left panel), or GST alone (GST, bottom right panel). Complexes were isolated and washed as described under “Experimental Procedures.” The samples were resolved by SDS-PAGE, and the gels were dried and processed by autoradiography. An aliquot of each [³⁵S]methionine-labeled TNT product (equivalent to 2% of the amount in each pulldown) was also resolved by SDS-PAGE and processed by autoradiography (Input, top left panel). The data are representative of five independent experiments.