Figure 2.
Influence of membrane composition on the solubilization of DHE by Osh4p. (A) Solubilization of DHE by Osh4p from DOPC liposomes doped with the indicated anionic lipid using the same protocol as in Fig. 1 E. When the fluorescence intensity was high enough, the I372/I340 ratio, which reflects the amount of DHE in complex with Osh4p in the supernatant (S) or in the pellet (P), was determined. (B) 0.5 µM NBD-Osh4p was added to buffer (−lip) or to liposomes (0.5 mM lipids) of various compositions. Trp fluorescence was monitored at 340 nm (λex = 285 nm; top), whereas NBD fluorescence was followed at 525 nm (λex = 495 nm; bottom). The contribution of the liposomes to the fluorescence signal was subtracted for each curve, and the fluorescence in buffer at t = 0 was used as a reference for normalization. Data shown in A are from a single representative experiment out of two independent experiments.