FIG 7 .

Identification of IQGAP1 as a novel binding partner of YopM^KIM. Ten micrograms of GST, GST-YopM^KIM, or the GST-YopM^KIM deletion variant indicated was loaded onto GST-Bind Sepharose beads and subsequently incubated with lysates prepared from LPS-primed C57BL/6 BMDMs or lysis buffer alone. Unbound proteins were removed, the beads were washed, and bound proteins were eluted and resolved by SDS-PAGE. (A) Silver staining detection of eluted proteins after incubation with BMDM lysate (lanes 1, 2, 4, and 5) or lysis buffer alone (lane 3). The arrow indicates an ~200-kDa protein that was excised from the gel, fragmented by trypsin digestion, subjected to analysis by LC-MS/MS, and identified as murine IQGAP1. (B) Western blot analysis of samples (starting material [SM], unbound [U], and bound [B]) from pulldown assays with BMDM lysate (lanes 1 to 5 and 8 to 11) or lysis buffer alone (lanes 6 and 7) with an antibody against IQGAP1 (188 kDa).