FIGURE 7.
FRET experiments using NBD-labeled FP5–13 and its derivatives serving as fluorescent donors, and Rho-labeled CP as an acceptor. A, spectra were obtained at room temperature with excitation set at 467 nm (10-nm slit) and an emission scan at 500–600 nm (10-nm slits). An NBD-labeled peptide was added to PC LUV (200 μm) in PBS to a final concentration of 0.4 μm peptide. This was followed by the addition of Rho-labeled CP peptide in several sequential doses. Fluorescence spectra were obtained first without Rho-peptide (upper curve) followed by four increasing Rho:NBD-labeled peptide ratios of 1:20 (second from top), 1:10 (third from top), and 1:7 (lower curve). B, nonlinear regression plot of FRET efficiency was determined by the relative percentage of emission at 530 nm between the NBD-labeled peptides alone and the peptides at different CP/NBD-labeled ratios.