Figure 5.

Ypt31-N126I and Ypt32-D129N inhibition of Ypt1 GEF. Purified GST-Bet3- or GST-containing fractions were tested for Ypt1-GEF activity using the GDP-release filtration assay with recombinant Ypt1 protein as described in the legend for Figure 1a. The GST-Bet3p reactions were carried out in the absence or presence of 1.6 μM Ypt31-N126I (●), 1.0 μM Ypt32-D129N (▴), or 1.6 μM Ypt31 wild-type (○) and 1.0 μM Ypt32 wild-type (▵) proteins. The results are expressed as percent inhibition by mutant or wild-type proteins of GEF-stimulated GDP-release. Intrinsic exchange, measured in the GST control, was subtracted from the values. Results shown in this figure are the average of duplicate measurements and are representative of two experiments. Error, calculated as average deviation (range divided by 2) was +/− 1% for all the data points.