Figure 3.

Specific interaction of bacterially expressed Ypt31/32-GTP with Myo2-GTD. His₆-tagged Ypt31 (pNS619), Ypt32 (pNS620), or Ypt1 (pNS1082) as a negative control, were expressed in bacteria and purified using Ni²⁺-NTA beads. The Ypts on the beads were loaded with GTPγS (T) or GDP (D) and incubated with lysates from bacteria expressing GST-tagged Myo2-GTD (pNS734), or GST (pNS255) as a negative control (left). The Ypts were precipitated using Ni²⁺ beads (right), and their presence was detected using immunoblot analysis and anti-His₆ antibodies (bottom). Coprecipitation of GST-Myo2-GTD or GST was detected using immunoblot analysis and anti-GST antibodies (top). GST-Myo2-GTD (∼5%), but not GST, coprecipitates with His₆-Ypt31 (A) and His₆-Ypt32 (B) preferentially in their GTP-bound form, but not with His₆-Ypt1. Results shown in this figure represent three independent experiments.