Functional sites of CFTR segregate to allosteric hotspots.

Shown is the amount of information (TE score) transmitted by each residue of dephosphorylated ATP-free human CFTR (PDB ID 5UAK) calculated using the ten most collective GNM modes (solid black trace). The positions of the 14 functionally important residues (see text for details) and of 30 ATP binding residues are shown as cyan and magenta spheres, respectively.

Identification of allosteric hotspots in dephosphorylated ATP-free CFTR.

A cartoon representation of dephosphorylated ATP-free human CFTR (PDB ID 5UAK), where each residue is colored according to the amount of information it transmits (TE score), expressed as Transfer Entropy x Collectivity (TECol) calculated using the ten most collective GNM modes (A), or upon removal of the first (B) or first and second (C) most collective GNM modes. Red and blue colors represent high and low levels of information output, respectively.

ATP binding and phosphorylation rewires and focuses the allosteric connectivity of CFTR.

(A) Shown is the amount of information transmitted (TE score) by each residue of phosphorylated ATP-bound human CFTR (PDB ID 6MSM) calculated using the ten most collective GNM modes (solid black trace). The positions of the ATP binding residues are shown as magenta spheres. (B,C) A cartoon representation of phosphorylated ATP-bound human CFTR (PDB ID 6MSM), where each residue is colored according to the amount of information it transmits (TE score), calculated using the ten most collective GNM modes (B), or upon removal of the first most collective GNM mode (C). Red and blue colors represent high and low levels of information output, respectively, and the ATP molecules are shown as black/grey spheres. (D) a magnified view of the allosteric cluster that is adjacent to the docking site of the R domain.

Synchronized movements of residues and domains of CFTR.

(A) Shown is a 2-D map of the dynamic cross-correlations between all residues of CFTR during the conformational transiting from the dephosphorylated ATP-free state to the phosphorylated ATP-bound state (PDB ID 5UAK and 6MSM, respectively). Red colors indicate strong positive correlation, meaning that the residues move in parallel vectors in space and time, blue colors indicate negative correlations. Domain boundaries are indicated by white lines. (B, C) Cartoon representation of CFTR where each residue is colored according to its dynamic cross- correlations with the ATP binding residues (black spheres) of NBD1 (B) or NBD2 (C).

Trajectory and sequence of allosteric transduction in CFTR.

Dynamic cross-correlations were calculated using a time delay (τ) of 16 cycles out of 50 required to complete the conformational transition between the dephosphorylated ATP-free and phosphorylated ATP-bound states. Red and blue colors indicate high and low correlations, respectively. Steps I-IV: Each residue is colored according to the degree of the correlation of its movement at time (t + τ) relative to the movement at time (t) of S1251 of NBD2 (Step 1), D110 of ECL1 (Step 2), T460 of NBD1 (Step 3), and the gating residue T338 (Step 4). Center (large) panel: shown is a summary of the allosteric trajectory, originating from NBD2 (green) to ICL2/3 and ECL1 (magenta), from ECL1 to NBD1 (blue), from NBD1 to the permeation pathway helices (orange), and finally back to NBD2.

Allosteric modulation of CFTR by drugs.

(A) Shown is the amount of information TRANSMITTED (TE score) by each residue of CFTR ΔF508 bound to Trikafta (PDB ID 8EIQ) calculated using the ten most collective GNM modes (solid black trace). The binding sites of ivacaftor, elexacaftor, and lumacaftor/tezacaftor are shown as magenta, green, or cyan spheres, respectively. (B) Cartoon representations of ivacaftor bound CFTR (PDB ID 6O2P), with ivacaftor and ATP shown as magenta and black/grey spheres, respectively. Residues are colored according to the amount of entropy RECEIVED from the ivacaftor binding residue F312 (C) Shown is the amount of entropy RECEIVED by each residue of CFTR from the ATP binding residue G551 (blue trace) or the ivacaftor binding residue F312 (orange trace). (D-F) Same as B, only the residues are colored according to the amount of entropy RECEIVED from the gating residue F337 (D), the ATP binding residue G551 (E), and the sum of the entropy received from both G551 and F337 (F).

Model of CFTR’s conformational changes and their allosteric control.

CFTR is shown in a grey space- filling model, the regulatory (R) domain as a red ribbon, and ATP in shades of green. Following its phosphorylation, the R domain dislodges from the NBDs dimer interface (Step I of the conformational change). This allows the ATP binding residues to fluctuate and sample conformers near the NBDs-open state (Step II), some of which provide an initial ATP binding site. In step III, this subset of conformers is selected by ATP to initiate binding (i.e., ensemble selection), which then proceeds to complete closure of the NBDs by an induced fit mechanism. In step IV, the channel bursts to the conducting state by a dynamic mechanism, without a major conformational change.