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289 J. Gen. Physiol. The Rockefeller University Press 0022-1295/97/03/289/11 $2.00 Volume 109 March 1997 289299
 

Summary: 289 J. Gen. Physiol. © The Rockefeller University Press · 0022-1295/97/03/289/11 $2.00
Volume 109 March 1997 289­299
Locating the Anion-selectivity Filter of the Cystic Fibrosis
Transmembrane Conductance Regulator (CFTR) Chloride Channel
Min Cheung* and Myles H. Akabas*§
From the *Center for Molecular Recognition, Department of Physiology and Cellular Biophysics, and §Department of Medicine, Col-
lege of Physicians and Surgeons, Columbia University, New York 10032
abstract The cystic fibrosis transmembrane conductance regulator forms an anion-selective channel; the site
and mechanism of charge selectivity is unknown. We previously reported that cysteines substituted, one at a time,
for Ile331, Leu333, Arg334, Lys335, Phe337, Ser341, Ile344, Arg347, Thr351, Arg352, and Gln353, in and flanking
the sixth membrane-spanning segment (M6), reacted with charged, sulfhydryl-specific, methanethiosulfonate
(MTS) reagents. We inferred that these residues are on the water-accessible surface of the protein and may line
the ion channel. We have now measured the voltage-dependence of the reaction rates of the MTS reagents with
the accessible, engineered cysteines. By comparing the reaction rates of negatively and positively charged MTS re-
agents with these cysteines, we measured the extent of anion selectivity from the extracellular end of the channel
to eight of the accessible residues. We show that the major site determining anion vs. cation selectivity is near the
cytoplasmic end of the channel; it favors anions by 25-fold and may involve the residues Arg347 and Arg352.
From the voltage dependence of the reaction rates, we calculated the electrical distance to the accessible residues.
For the residues from Leu333 to Ser341 the electrical distance is not significantly different than zero; it is signifi-
cantly different than zero for the residues Thr351 to Gln353. The maximum electrical distance measured was 0.6

  

Source: Akabas, Myles - Department of Physiology and Biophysics, Albert Einstein College of Medicine, Yeshiva University

 

Collections: Biology and Medicine