Figure 3.

 IAA Binds to SKP2A, but Not to SKP2B.

(A) Pull-down reactions were performed with either MBP-SKP2A or MBP-SKP2B bound to amylase beads in the presence of 50 nM [³H]-IAA. The retained [³H]-IAA in the amylose beads after three washes was measured by scintillation counting. Each value is the mean of three independent measures, and the error bars correspond to the sd. d.p.m., disintegrations per minute.

(B) Competitive binding of [³H]-IAA to MBP-SKPA and MBP-SKP2B in the presence of increasing concentrations of unlabeled IAA. Pull-down reactions were performed in the presence of 50 nM [³H]-IAA and the indicated amount of competitor. Each value corresponds to the mean of three independent experiments. Error bars represent the sd.

(C) Competitive binding of [³H]-IAA to MBP-SKPA in the presence of increasing concentrations of unlabeled 1-NAA (black squares), 2-NAA (gray squares), or 2,4-D (black triangles). MBP alone was used as a control of nonbinding protein. Pull-down reactions were performed in the presence of 50 nM [³H]-IAA and the indicated amount of cold competitor. The inactive auxin 2-NAA does not compete with [³H]-IAA. Values are the mean of three independent experiments. Error bars represent the sd.