Fig. 3: CFTR allosteric dynamics perturbation by deletion of the F508 sidechain.
a Root mean square fluctuations (RMSF) are indicated on the CFTR CL4 structure using color and thickness coding. Higher RMSF values are indicated by warmer colors and thicker tube and calculated from n = 6 MD trajectories (frames = 30,000). b Fraction of native contacts of CL4 and CL1 with the rest of the protein, and between NBD1 and NBD2 were calculated over all trajectories and plotted for WT- and F508G-CFTR, as well as F508G-CFTR-6SS. c Allosteric coupling was investigated by community analysis of residue networks based on motion correlation. Amino acids in a community are color-coded. The spheres indicate the community size. The thickness (1-weight) of edges between spheres represents the dynamic coupling between communities. Black dotted and solid ovals highlight the alterations in coupling between the α-subdomain of NBD1 and NBD2 in WT (weight between communities: 0.6576), F508G (0.6922), and F508G-6SS (0.9274), and between the distant TMH4/5(CL2) and Lasso in WT (0.6576) or F508G (0.6918), and F508G-6SS (0.9285), respectively. d The Tm of the isolated NBD1 variants of CFTR, MRP1, and ABCC6. Means ± S.E.M., n = 4,3,6,6,3,3 from right to left. Biological replicates. e The folding free energy (ΔGo) and unfolding activation energy (ΔG#u) of WT and mutant NBD1 variants of ABCC6 and MRP1 were estimated by CD-spectroscopy with urea- and temperature-dependent unfolding as described in Methods. ΔGo: means ± S.D.M., n = 3, ΔG#u, extrapolated values based on n = 2 experiments at 4 different temperatures. f, g Effect of NBD1-CL7 interface uncoupling on the complex- (C-band) and core-glycosylated (B-band) ABCC6 and MRP1 expression in BHK-21 cells (f), determined by quantitative immunoblotting (g). Only 10 or 20% of WT variants were analyzed. Means ± S.E.M., n = 4. Biological replicates. h The metabolic stabilities of complex-glycosylated MRP1 and ABCC6 were monitored by CHX chase and quantitative immunoblotting. Means ± S.E.M., n = 3 except WT-ABCC6; n = 5. Biological replicates. WT:red, mutants:green. Source data are provided in Source Data file.
