FIGURE 5.

The effect of chemical correction and ablation of the R⁵⁵³XR⁵⁵⁵ sequence on conformational correction of F508del-CFTR probed using limited proteolysis. Limited proteolysis of control or chemically corrected F508del-CFTR, F508del-KXK-CFTR, or WT-CFTR with increasing levels of trypsin (0, 1.56, 3.13, 6.25, 12.5, 25, 50, 100 μg/ml) was performed. Digests were analyzed by SDS-PAGE, and immunoreactive bands were detected using the Odyssey infrared imaging system from LI-COR Biosciences. The colored signal was converted to grayscale for the preparation of this figure. A, samples were immunoblotted with anti-CFTR monoclonal antibody, L12B4 (NBD1-specific). Subsequent analyses focused on the larger NBD1 fragments (34–36 kDa) and the smaller NBD1 fragments (lower than 28 kDa). Both sets of fragments were indicated using brackets on the right-hand side of each image. B, samples were immunoblotted with anti-CFTR monoclonal antibody, M3A7 (NBD2-specific). NBD2 fragments of ∼28 kDa were analyzed subsequently (indicated using brackets on the right-hand side of each image).