FIGURE 3.

Fluorescence intensity of NBD attached to cysteine-substituted LC-HCT in soluble and liposome-associated forms. Emission scans are shown of the soluble (red lines) and liposome-bound (black lines) forms of I830C-NBD (A) and I684C-NBD (B) at pH 4.4. The large increase in intensity and blue shift to 530 nm when I830C-NBD bound to liposomes is consistent with movement of NBD into a non-polar environment. C, compilation of the NBD intensity changes (F_mem/F_sol) observed at 530 nm for specific LC-HCT residues when proteins in the soluble (F_sol) and liposome-associated (F_mem) forms are compared. Residues were selected throughout the HCT structure with an emphasis on the protease-protected region identified in Fig. 2 (residues 805–836; green). Residues from the neighboring loop (sequence 682–688; orange) are included to show that this change is specific to the protease-protected region. D, close-up of the location of tested residues within the context of the LC-HCT structure. Residues with F_mem/F_sol > 3 are shown in blue, and residues with F_mem/F_sol < 3 are shown in red.