FIGURE 8.

Microsomal Slc1p activity is protease sensitive. Microsomes (micros.) from ale1Δ cells (50 μg of protein/sample) were subject to three consecutive preincubations at 0 °C lasting 60, 60, and 10 min (right), with the compounds listed on the left (added in the order from top down). First, for protease pretreatments, microsomes were incubated with or without Triton X-100 (32 μg = 51 nmol) and trypsin (200 μg). Second, protease inhibitors (Inhib.; antipain and aprotinin, 200 μg each) were added. Third, Triton X-100 and/or a second aliquot of microsomes were added to some samples. In samples 11–14, S-NHS (400 nmol) was added instead of trypsin and later inactivated by adding l-lysine (120 μmol). Subsequent to the three indicated preincubations, the LPAAT activity was assayed at 0 °C for 30 s as described under “Experimental Procedures.” For sample 2, boiled microsomes were used. The plot reports mean cpm as detected by Berthold radioscanning of two independent experiments. Error bars, S.E.