Figure 8.

Sec2-59p–stimulated guanine nucleotide exchange on Sec4p. (A) Aliquots containing 1 μM Sec4p-[³H]GDP were incubated with the indicated levels of Sec2-59p or buffer in the presence of excess unlabeled GDP. At the time intervals indicated, Sec4p-bound radioactivity was determined by the filter-binding assay. To retard the intrinsic off rate of Sec4p, this experiment was performed at 14°C. (B) Aliquots containing 1 μM Sec4p were incubated with Sec2-59p or buffer, as indicated in the presence of [³⁵S]GTPγS at 14°C. At the time intervals indicated, Sec4p-bound radioactivity was determined by the filter-binding assay. For each assay, the relative values correspond to the nanomoles of GTPγS retained on the filter.