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Figure 2—figure supplement 1. Non-native inter-NBD H-bond in OF E-to-Q mutant hCFTR, but not zCFTR, suggested by cryo-EM structures. (A) Macroscopic current relaxations upon ATP removal in inside-out patches excised from Xenopus laevis oocytes expressing hE1371Q (black trace), hE1371S (dark blue trace), zE1372Q (brown trace), and zE1372S (light blue trace) CFTR channels. Currents were activated by exposure to saturating ATP, following prior phosphorylation for ~2 min with 300 nM bovine PKA catalytic subunit. Current amplitudes are shown normalized by their steady-state values in ATP (i.e., just before ATP removal), membrane potential (Vm) was −20 to −80 mV. (B) Mean burst durations (in seconds) obtained as the time constants of single-exponential fits to the macroscopic relaxations shown in (A). Data are shown as mean ± standard error of the mean (SEM) from six to nine experiments using a logarithmic ordinate. (A, B) has been adapted from Figures 2A, B, and 3C, D from Simon and Csanády, 2023. (C, D) Cryo-EM structures of hCFTR-E1371Q (PDBID: 6msm) and zCFTR-E1372Q (PDBID: 5w81) in the OF conformation. Color coding as in (A), positions hG576/zT575 (orange), hQ1371 (black), and zQ1372 (brown) are shown in spacefill. Red dotted squares identify regions expanded in panels (E, F). (E, F) Close-up views of the regions surrounding the mutated catalytic glutamate side chains (black/brown sticks) in OF hCFTR (E) and zCFTR (F).