Cystic fibrosis transmembrane conductance regulator (CFTR) anion binding as a probe of the pore.
We compared the effects of mutations in transmembrane segments (TMs) TM1, TM5, and TM6 on the conduction and activation properties of the cystic fibrosis transmembrane conductance regulator (CFTR) to determine which functional property was most sensitive to mutations and, thereby, to develop a criterion for measuring the importance of a particular residue or TM for anion conduction or activation. Anion substitution studies provided strong evidence for the binding of permeant anions in the pore. Anion binding was highly sensitive to point mutations in TM5 and TM6. Permeability ratios, in contrast, were relatively unaffected by the same mutations, so that anion binding emerged as the conduction property most sensitive to structural changes in CFTR. The relative insensitivity of permeability ratios to CFTR mutations was in accord with the notion that anion-water interactions are important determinants of permeability selectivity. By the criterion of anion binding, TM5 and TM6 were judged to be likely to contribute to the structure of the anion-selective pore, whereas TM1 was judged to be less important. Mutations in TM5 and TM6 also dramatically reduced the sensitivity of CFTR to activation by 3-isobutyl 1-methyl xanthine (IBMX), as expected if these TMs are intimately involved in the physical process that opens and closes the channel.
Mansoura
M K
MK
Department of Biomedical Engineering, University of Michigan, Ann Arbor 48109-0622, USA.
Smith
S S
SS
Choi
A D
AD
Richards
N W
NW
Strong
T V
TV
Drumm
M L
ML
Collins
F S
FS
Dawson
D C
DC
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D019943:Amino Acid Substitution; D000818:Animals; D000838:Anions / Q000378:metabolism; D001665:Binding Sites; D002463:Cell Membrane Permeability; D000242:Cyclic AMP / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D004553:Electric Conductivity; D005260:Female; D005998:Glycine; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000502:physiology*; D017354:Point Mutation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013861:Thiocyanates / Q000494:pharmacology; D014982:Xenopus laevis
D000838:Anions; C505032:CFTR protein, human; D011994:Recombinant Proteins; D013861:Thiocyanates; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000242:Cyclic AMP; D015056:1-Methyl-3-isobutylxanthine; D005998:Glycine