Figure 2.

Early-import intermediate formation is reduced when GTP/GDP exchange is inhibited. Chloroplasts were prepared as described in Figure 1. Chloroplasts were then incubated with a 1 mm analog concentration with equimolar Mg²⁺ for 5 min. Radiolabeled prSS and 0, 0.1, or 1.0 mm MgATP was added and incubated for 30 min. For the GTP/GDP competition studies (lanes 7–9), 500 μm GTP and GDP were added to chloroplasts for 5 min before the addition of MgATP. Samples were analyzed by SDS-PAGE and fluorography (A) and were quantitated with a phosphor imager (B and C). The control for quantitation of intermediate formation was lane 2 of A (prSS bound in the presence of 0.1 mm ATP), whereas the control for translocation was lane 3 of A (prSS translocated in the presence of 1 mm ATP). TP, Translated product (10%) added to a single reaction. Results shown are from one of three separate experiments.