FIGURE 1.

Identification of IQGAP1 as a novel S100P target protein. A, affinity chromatography of placental proteins on columns containing immobilized S100Pwt or the monomeric F15A mutant. Equal amounts of placental protein extract were loaded onto the columns in a Ca²⁺-containing buffer, and flow-through fractions were collected (FT). Columns were washed, (W) and Ca²⁺-dependently bound proteins were eluted with an EGTA-containing buffer (E). The collected fractions were subjected to SDS-PAGE and stained with Coomassie. MALDI-TOF analysis identified the protein migrating at an apparent molecular mass of 190 kDa as human IQGAP1. This protein was only present in the eluate of the S100Pwt column. B, affinity chromatography with S100P and IQGAP1 reveals that the interaction is direct and Ca²⁺-dependent. S100Pwt was loaded onto columns containing immobilized GST-IQGAP1 or GST alone in a Ca²⁺- or EGTA-containing buffer, and flow-through fractions were collected (FT). Columns were washed (W) and Ca²⁺-dependently bound S100Pwt was eluted with an EGTA-containing buffer (E). The collected fractions were analyzed by immunoblotting for S100P.