Figure 5.

Toc34 interaction with preSSU is regulated by phosphorylation and a GTP/GDP cycle. A total of 20 pM [³²P]preSSU was incubated with 10 nM Toc34ΔTM252–6His (lanes 1–4) and 1 μM ATP in the presence (lanes 3 and 4) or absence (lanes 1 and 2) of 1 mM GTP, and in the presence (lanes 2 and 4) or absence (lanes 1 and 3) of a protein kinase-enriched wheat germ fraction (0.5 ng total protein, KWGF); complex formation was tested by immunoprecipitation. (B) A total of 20 pM of [³²P]preSSU was incubated with 10 nM Toc34ΔTM252–6His in the presence of 1 mM GTP (lane 1). After 10 min of preincubation, 10 mM of ADP (lane 2) or GDP (lane 3) was added, and the remaining complex was immunoprecipitated. Histograms represent averaged value of three independent experiments. (A) Binding to nonphosphorylated Toc34 was set to 100%; (B) binding without competition was set to 100%.