Allosteric Modulation Balances Thermodynamic Stability and Restores Function of ΔF508 CFTR

JOURNAL OF MOLECULAR BIOLOGY

卷:419

Article

Aleksandrov, Andrei A.^1,2  Kota, Pradeep^1,3  Cui, Liying^1,2  Jensen, Tim^1,2  Alekseev, Alexey E.⁴  Reyes, Santiago⁴  He, Lihua^1,2  Gentzsch, Martina^2,5  Aleksandrov, Luba A.^1,2  Dokholyan, Nikolay V.^1,3  Riordan, John R.^1,2 

[1] Univ N Carolina, Dept Biochem & Biophys, Chapel Hill, NC 27599 USA

[2] Univ N Carolina, Cyst Fibrosis Treatment & Res Ctr, Chapel Hill, NC 27599 USA

[3] Univ N Carolina, Mol & Cellular Biophys Program, Chapel Hill, NC 27599 USA

[4] Mayo Clin, Div Cardiovasc Dis, Dept Med, Rochester, MN 55905 USA

[5] Univ N Carolina, Dept Cell & Dev Biol, Chapel Hill, NC 27599 USA

关键词: ABC transporters;    CFTR;    protein thermal stability;    ion channel;    DMD simulations;   

DOI  :  10.1016/j.jmb.2012.03.001

来源: Elsevier

【 摘 要 】

Most cystic fibrosis is caused by a deletion of a single residue (F508) in CFTR (cystic fibrosis transmembrane conductance regulator) that disrupts the folding and biosynthetic maturation of the ion channel protein. Progress towards understanding the underlying mechanisms and overcoming the defect remains incomplete. Here, we show that the thermal instability of human Delta F508 CFTR channel activity evident in both cell-attached membrane patches and planar phospholipid bilayers is not observed in corresponding mutant CETRs of several non-mammalian species. These more stable orthologs are distinguished from their mammalian counterparts by the substitution of proline residues at several key dynamic locations in first N-terminal nucleotide-binding domain (NBD1), including the structurally diverse region, the gamma-phosphate switch loop, and the regulatory insertion. Molecular dynamics analyses revealed that addition of the prolines could reduce flexibility at these locations and increase the temperatures of unfolding transitions of Delta F508 NBD1 to that of the wild type. Introduction of these prolines experimentally into full-length human Delta F508 CFTR together with the already recognized I539T suppressor mutation, also in the structurally diverse region, restored channel function thermodynamic stability as well as its trafficking to and lifetime at the cell surface. Thus, while cellular manipulations that circumvent its culling by quality control systems leave Delta F508 CFTR dysfunctional at physiological temperature, restoration of the delicate balance between the dynamic protein's inherent stability and channel activity returns a near-normal state. (C) 2012 Elsevier Ltd. All rights reserved.

【 授权许可】

Free