Figure 1.

GTP has a separate role that is distinct from the ATP requirement during the generation of early-import intermediates. Chloroplasts were depleted of endogenous levels of ATP by incubation with nigericin for 10 min in the dark. They were then incubated with a 1 mm concentration of GTP analog with equimolar Mg²⁺ for 5 min (except for the experiment shown in lane 16, in which a 5 mm analog concentration was used). Radiolabeled prSS and 0, 0.1, or 1.0 mm MgATP was added and incubated for 30 min. 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.