Molecular models of the open and closed states of the whole human CFTR protein

Mornon, Jean‐Paul; Lehn, Pierre; Callebaut, Isabelle

doi:10.1007/s00018-009-0133-0

Cited by 142 publications

(241 citation statements)

References 70 publications

(125 reference statements)

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“…One possibility is that each corrector interacts with a different structural feature of F508del-CFTR to stabilize their folding or reduce nonproductive folding intermediates. Because the folding defect of F508del involves multiple domain and interdomain interactions, multiple interaction sites may be necessary for optimal correction of the misfolded protein (33)(34)(35)(36)(37). Alternatively, rescue might be achieved indirectly through the modulation of folding chaperones or by suppressing ER-associated degradation (ERAD) quality control pathways that promote F508del-CFTR ubiquitylation and degradation.…”

Section: Discussionmentioning

confidence: 99%

Pharmacological Rescue of the Mutant Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Detected by Use of a Novel Fluorescence Platform

Holleran

Glover

Peters

et al. 2012

Mol Med

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Numerous human diseases arise because of defects in protein folding, leading to their degradation in the endoplasmic reticulum. Among them is cystic fibrosis (CF), caused by mutations in the gene encoding the CF transmembrane conductance regulator (CFTR ), an epithelial anion channel. The most common mutation, F508del, disrupts CFTR folding, which blocks its trafficking to the plasma membrane. We developed a fluorescence detection platform using fluorogen-activating proteins (FAPs) to directly detect FAP-CFTR trafficking to the cell surface using a cell-impermeant probe. By using this approach, we determined the efficacy of new corrector compounds, both alone and in combination, to rescue F508del-CFTR to the plasma membrane. Combinations of correctors produced additive or synergistic effects, improving the density of mutant CFTR at the cell surface up to ninefold over a single-compound treatment. The results correlated closely with assays of stimulated anion transport performed in polarized human bronchial epithelia that endogenously express F508del-CFTR. These findings indicate that the FAP-tagged constructs faithfully report mutant CFTR correction activity and that this approach should be useful as a screening assay in diseases that impair protein trafficking to the cell surface.

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Section: Discussionmentioning

confidence: 99%

Pharmacological Rescue of the Mutant Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Detected by Use of a Novel Fluorescence Platform

Holleran

Glover

Peters

et al. 2012

Mol Med

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“…EM data (29-31) do not provide detailed information on interactions with R region, making it challenging to address the structural implications of phosphorylation on the full-length protein. In addition, there is debate regarding the position of R region with respect to other domains of CFTR based on modeling (32,33). Using our NMR binding information obtained on isolated domains of CFTR, we performed computational modeling to provide a structural view of full-length CFTR in the nonphosphorylated state.…”

Section: R Region Binds Multiple Partners In a Phosphorylation-dependentmentioning

confidence: 99%

Regulatory R region of the CFTR chloride channel is a dynamic integrator of phospho-dependent intra- and intermolecular interactions

Bozóky

Krzeminski

Muhandiram

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

151

218

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Intrinsically disordered proteins play crucial roles in regulatory processes and often function as protein interaction hubs. Here, we present a detailed characterization of a full-length disordered hub protein region involved in multiple dynamic complexes. We performed NMR, CD, and fluorescence binding studies on the nonphosphorylated and highly PKA-phosphorylated human cystic fibrosis transmembrane conductance regulator (CFTR) regulatory region, a ∼200-residue disordered segment involved in phosphorylation-dependent regulation of channel trafficking and gating. Our data provide evidence for dynamic, phosphorylation-dependent, multisite interactions of various segments of the regulatory region for its intra-and intermolecular partners, including the CFTR nucleotide binding domains 1 and 2, a 42-residue peptide from the C terminus of CFTR, the SLC26A3 sulphate transporter and antisigma factor antagonist (STAS) domain, and 14-3-3β. Because of its large number of binding partners, multivalent binding of individually weak sites facilitates rapid exchange between free and bound states to allow the regulatory region to engage with different partners and generate a graded or rheostat-like response to phosphorylation. Our results enrich the understanding of how disordered binding segments interact with multiple targets. We present structural models consistent with our data that illustrate this dynamic aspect of phospho-regulation of CFTR by the disordered regulatory region.protein interaction network | fuzzy complex

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“…The outward facing models 14,[17][18][19][20][21][22][23] were based primarily on the Sav1866 structure as a template, 24 although recently the newly solved ABC transporter structure McjD 25 has been used to model this state. 26 Inward facing models were based either on the bacterial MsbA 27 or on the P-gp 28 structures.…”

Section: Introduction Cystic Fibrosis (Cf) Is a Lethal Inherited Dismentioning

confidence: 99%

Molecular Dynamics Flexible Fitting Simulations Identify New Models of the Closed State of the Cystic Fibrosis Transmembrane Conductance Regulator Protein

Simhaev

McCarty

Ford

et al. 2017

J. Chem. Inf. Model.

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Cystic Fibrosis (CF) is a lethal, genetic disease found in particular in humans of European origin which is caused by mutations in the CFTR chloride channel. The search for CF therapies acting by modulating the impaired function of mutant CFTR will be greatly advanced by high resolution structures of CFTR in different states. To date, two medium resolution EM structures of CFTR are available (one of a distant zebrafish (Danio rerio) CFTR ortholog and one of human CFTR). The two models are nearly identical to one another and both correspond to the inward-facing, NBDs separated, closed state of the channel. In addition, lower resolution structural data are available for human CFTR in an alternative conformation which likely features associated NBDs and thus geometrically resembles the conducting state of the channel.Multiple homology models of human CFTR in multiple states have been developed over the years, yet their correspondence to the existing structural information is unexplored. In this work we use molecular dynamics flexible fitting (MDFF) simulations to refine two previously described CFTR models based on the available cryo-EM map of the human protein. This map was recorded in the absence of ATP and consequently represents closed-state CFTR yet its features likely correspond to a NBDs associated conformation of the protein. Accordingly, the resulting models feature dimerized NBDs yet with no membrane traversing pore. Moreover, the open probability of the new models as deduced from the MDFF trajectories is significantly lower than that deduced from control MD trajectories initiated from the starting models. We propose that the new models correspond to a CFTR conformation which to date was largely unexplored yet one that is relevant to the gating cycle of the protein. In particular this conformation may participate in rapid channel opening and closing through small allosteric movements controlled by nucleotide binding and dissociation events. Analyzing the resulting trajectories (and not only the final models as is usually the case), we demonstrate that the refined models have good stereochemical properties and are also in favorable agreement with multiple experimental data.Moreover, despite different starting points, the final models share many common features.Finally, we propose that the combination of high resolution cryo-EM maps which are currently emerging from multiple labs and MDFF simulations will be of value for the development of yet more reliable CFTR models as well as for the identification of binding sites for CFTR modulators.

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Molecular models of the open and closed states of the whole human CFTR protein

Cited by 142 publications

References 70 publications

Pharmacological Rescue of the Mutant Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Detected by Use of a Novel Fluorescence Platform

Pharmacological Rescue of the Mutant Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Detected by Use of a Novel Fluorescence Platform

Regulatory R region of the CFTR chloride channel is a dynamic integrator of phospho-dependent intra- and intermolecular interactions

Molecular Dynamics Flexible Fitting Simulations Identify New Models of the Closed State of the Cystic Fibrosis Transmembrane Conductance Regulator Protein

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