Véronique Stoven

A novel model for the first nucleotide binding domain of the cystic fibrosis transmembrane conductance regulator

Cystic fibrosis is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. The most frequent mutation is the deletion of F508 in the first nucleotide binding fold (NBF1). It induces a perturbation in the folding of NBF1, which impedes posttranslational maturation of CFTR. Determination of the three‐dimensional structure of NBF1 would help to understand this defect. We present a novel model for NBF1 built from the crystal structure of bovine mitochondrial F1‐ATPase protein. This model gives a reasonable interpretation of the effect of mutations on the maturation of the protein and, in agreement with the CD data, leads to reconsideration of the limits of NBF1 within CFTR.

A New N-Channel Maximum Entropy Method in NMR for Automatic Reconstruction of “Decoupled Spectra” and J-Coupling Determination

A new N-channel (or multichannel) deconvolution method of 1D and 2D NMR spectra is presented based on the Maximum Entropy method. In the general case of spectra exhibiting several lines of various fine structures, the method allows simultaneous deconvolution of all lines. While deconvolution is performed, determination of the values of the coupling constants is possible. Reconstruction of “decoupled” 1D and 2D spectra is also performed in a fully automated approach. The efficiency of the method is illustrated on experimental 1D and 2D spectra. The method has been developed in the GIFA software, a complete package for processing of NMR data available from the authors.

Insight into cystic fibrosis by structural modelling of CFTR first nucleotide binding fold (NBF1).

Cystic fibrosis is a human monogenic genetic disease caused by mutations in the cystic fibrosis (CF) gene, which encodes a membrane protein which functions as a channel: the cystic fibrosis transmembrane conductance regulator (CFTR) protein. The most frequent mutation, a deletion of phenylalanine F508 (delta F508), is located in the first nucleotide binding domain of CFTR: NBF1. This mutation leads to a folding defect in NBF1, responsible for an incomplete maturation of CFTR. The absence of CFTR at the surface of epithelial cells causes the disease. Determination of the three-dimensional (3D) structure of NBF1 is a key step to understanding the alterations induced by the mutation. In the absence of any experimental data, we have chosen to build a 3D model for NBF1. This model was built by homology modelling starting from F1-ATPase, the only protein of known 3D structure in the ATP binding cassette (ABC) family. This new model defines the central and critical position of F508, predicted in the hydrophobic core of NBF1. F508 indeed could be involved in hydrophobic interactions to ensure a correct folding pathway. Moreover, this model enables the localization of the LSGGQ sequence (a highly conserved sequence in the ABC family) in a loop, at the surface of the protein. This reinforces the hypothesis of its role for mediation of domain-domain interactions of functional significance for the channel regulation. Finally, the model also allows redefinition of the ends of NBF1 within the CFTR sequence. These extremities are defined by the secondary structure elements that are involved in the NBF1 fold. They lead to reconsideration of the C-terminal limit which was initially defined by the end of exon 12.