RT Journal Article
SR Electronic
T1 A Small-Molecule Modulator Interacts Directly with ΔPhe508-CFTR to Modify Its ATPase Activity and Conformational Stability
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 1430
OP 1438
DO 10.1124/mol.109.055608
VO 75
IS 6
A1 Leigh Wellhauser
A1 Patrick Kim Chiaw
A1 Stan Pasyk
A1 Canhui Li
A1 Mohabir Ramjeesingh
A1 Christine E. Bear
YR 2009
UL http://molpharm.aspetjournals.org/content/75/6/1430.abstract
AB The deletion of Phe-508 (ΔPhe508) constitutes the most prevalent of a number of mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) that cause cystic fibrosis (CF). This mutation leads to CFTR misfolding and retention in the endoplasmic reticulum, as well as impaired channel activity. The biosynthetic defect can be partially overcome by small-molecule “correctors”; once at the cell surface, small-molecule “potentiators” enhance the channel activity of ΔPhe508-CFTR. Certain compounds, such as VRT-532, exhibit both corrector and potentiator functions. In the current studies, we confirmed that the inherent chloride channel activity of ΔPhe508-CFTR (after biosynthetic rescue) is potentiated in studies of intact cells and membrane vesicles. It is noteworthy that we showed that the ATPase activity of the purified and reconstituted mutant protein is directly modulated by binding of VRT-532 [4-methyl-2-(5-phenyl-1H-pyrazol-3-yl)-phenol] ATP turnover by reconstituted ΔPhe508-CFTR is decreased by VRT-532 treatment, an effect that may account for the increase in channel open time induced by this compound. To determine whether the modification of ΔPhe508-CFTR function caused by direct VRT-532 binding is associated with structural changes, we evaluated the effect of VRT-532 binding on the protease susceptibility of the major mutant. We found that binding of VRT-532 to ΔPhe508-CFTR led to a minor but significant decrease in the trypsin susceptibility of the full-length mutant protein and a fragment encompassing the second half of the protein. These findings suggest that direct binding of this small molecule induces and/or stabilizes a structure that promotes the channel open state and may underlie its efficacy as a corrector of ΔPhe508-CFTR. The American Society for Pharmacology and Experimental Therapeutics