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The ΔF508 cystic fibrosis mutation impairs domain-domain interactions and arrests post-translational folding of CFTR

Nature Structural & Molecular Biology volume 12, pages 1725 (2005) | Download Citation

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Abstract

Misfolding accounts for the endoplasmic reticulum–associated degradation of mutant cystic fibrosis transmembrane conductance regulators (CFTRs), including deletion of Phe508 (ΔF508) in the nucleotide-binding domain 1 (NBD1). To study the role of Phe508, the de novo folding and stability of NBD1, NBD2 and CFTR were compared in conjunction with mutagenesis of Phe508. ΔF508 and amino acid replacements that prevented CFTR folding disrupted the NBD2 fold and its native interaction with NBD1. ΔF508 caused limited alteration in NBD1 conformation. Whereas nonpolar and some aliphatic residues were permissive, charged residues and glycine compromised the post-translational folding and stability of NBD2 and CFTR. The results suggest that hydrophobic side chain interactions of Phe508 are required for vectorial folding of NBD2 and the domain-domain assembly of CFTR, representing a combined co- and post-translational folding mechanism that may be used by other multidomain membrane proteins.

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Acknowledgements

We are indebted to N. Kartner and D. Bedwell for generously providing M3A7, L12B4 and antibody 4562 anti-CFTRs. We thank C. Daniels for careful reading the manuscript. M.S. was supported by a Canadian Institutes of Health Research (CIHR) doctoral studentship. This work was supported by grants to G.L. from the Canadian Cystic Fibrosis Foundation, the CIHR and the Premier's Research Excellence Award of the Ontario Ministry of Energy and Education.

Author information

Affiliations

  1. Hospital for Sick Children Research Institute, Program in Cell and Lung Biology, University of Toronto, Ontario M5G 1X8, Canada.

    • Kai Du
    • , Manu Sharma
    •  & Gergely L Lukacs
  2. Department of Laboratory Medicine and Pathobiology, University of Toronto, Ontario M5G 1X8, Canada.

    • Manu Sharma
    •  & Gergely L Lukacs

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Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Gergely L Lukacs.

Supplementary information

PDF files

  1. 1.

    Supplementary Fig. 1

    NBD1 fragment recognized by two antibodies.

  2. 2.

    Supplementary Fig. 2

    Conformational maturation of NBDs.

  3. 3.

    Supplementary Fig. 3

    Anion channel function of X508 CFTR mutants.

  4. 4.

    Supplementary Fig. 4

    Proteinase K susceptibility of NBD2.

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DOI

https://doi.org/10.1038/nsmb882

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