title: Evidence for direct CFTR inhibition by CFTR(inh)-172 based on Arg347 mutagenesis.
abstract: CFTR (cystic fibrosis transmembrane conductance regulator) is an epithelial Cl- channel inhibited with high affinity and selectivity by the thiazolidinone compound CFTR(inh)-172. In the present study, we provide evidence that CFTR(inh)-172 acts directly on the CFTR. We introduced mutations in amino acid residues of the sixth transmembrane helix of the CFTR protein, a domain that has an important role in the formation of the channel pore. Basic and hydrophilic amino acids at positions 334-352 were replaced with alanine residues and the sensitivity to CFTR(inh)-172 was assessed using functional assays. We found that an arginine-to-alanine change at position 347 reduced the inhibitory potency of CFTR(inh)-172 by 20-30-fold. Mutagenesis of Arg347 to other amino acids also decreased the inhibitory potency, with aspartate producing near total loss of CFTR(inh)-172 activity. The results of the present study provide evidence that CFTR(inh)-172 interacts directly with CFTR, and that Arg347 is important for the interaction.
authors: Laboratorio di Genetica Molecolare, Istituto Giannina Gaslini, L.go Gerolamo Gaslini 5, 16147 Genova, Italy.
authors: Emanuela
authors: E
authors: Caci
authors: Antonella
authors: A
authors: Caputo
authors: Alexandre
authors: A
authors: Hinzpeter
authors: Nicole
authors: N
authors: Arous
authors: Pascale
authors: P
authors: Fanen
authors: Nitin
authors: N
authors: Sonawane
authors: A S
authors: AS
authors: Verkman
authors: Roberto
authors: R
authors: Ravazzolo
authors: Olga
authors: O
authors: Zegarra-Moran
authors: Luis J V
authors: LJ
authors: Galietta
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001120:Arginine; D001565:Benzoates / Q000737:chemistry / Q000494:pharmacology*; D002460:Cell Line; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000037:antagonists & inhibitors*; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology; D015394:Molecular Structure; D016296:Mutagenesis; D011485:Protein Binding; D051381:Rats; D053778:Thiazolidines / Q000737:chemistry / Q000494:pharmacology*
chemical_list: C482900:3-((3-trifluoromethyl)phenyl)-5-((3-carboxyphenyl)methylene)-2-thioxo-4-thiazolidinone; D001565:Benzoates; D053778:Thiazolidines; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D001120:Arginine