Fig. 4. Gyp7p inhibition is mediated via GTP hydrolysis. Fusion reactions (150 µl) were incubated at 27°C with Gyp7p, GTPγS, or both Gyp7p and GTPγS. After 30 min, vacuoles were re-isolated and aliquots added to reaction buffer with no inhibitor or with Gyp7p, Gdi1p, anti-Vam3p antibodies or GTPγS, and incubated for 60 min at 27°C before being assayed for alkaline phosphatase.