Channel-lining residues in the M3 membrane-spanning segment of the cystic fibrosis transmembrane conductance regulator

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Abstract

The cystic fibrosis transmembrane conductance regulator (CFTR) forms a chloride-selective channel. Residues from the 12 putative membrane-spanning segments form at least part of the channel lining. We need to identify the channel-lining residues in order to understand the structural basis for the channel's functional properties. Using the substituted-cysteine- accessibility method we mutated to cysteine, one at a time, 24 consecutive residues (Asp192-Ile215) in the M3 membrane-spanning segment. Cysteines substituted for His 199, Phe200, Trp202, Ile203, Pro205, Gln207, Leu211, and Leu214 reacted with charged, sulfhydryl-specific reagents that are derivatives of methanethiosulfonate (MTS). We infer that these residues are on the water-accessible surface of the protein and probably form a portion of the channel lining. When plotted on an α-helical wheel the exposed residues from Gln207 to Leu214 lie within an arc of 60°; the exposed residues in the cytoplasmic half (His199-Ile203) lie within an arc of 160°. We infer that the secondary structures of the extracellular and cytoplasmic halves of M3 are α-helical and that Pro205, in the middle of the M3 segment, may bend the M3 segment, moving the cytoplasmic end of the segment in toward the central axis of the channel. The bend in the M3 segment may help to narrow the channel lumen near the cytoplasmic end. In addition, unlike full-length CFTR, the current induced by the deletion construct, Δ259, is inhibited by the MTS reagents, implying that the channel structure of Δ259 is different than the channel structure of wild-type CFTR.

Original languageEnglish (US)
Pages (from-to)12233-12240
Number of pages8
JournalBiochemistry
Volume37
Issue number35
DOIs
StatePublished - Sep 1 1998
Externally publishedYes

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Cystic Fibrosis Transmembrane Conductance Regulator
Linings
Cysteine
Membranes
Cytoplasmic Structures
Sulfhydryl Reagents
Chloride Channels
Induced currents
Chlorides
Wheels
Membrane Proteins
Derivatives
Water
methanethiosulfonate

ASJC Scopus subject areas

  • Biochemistry

Cite this

Channel-lining residues in the M3 membrane-spanning segment of the cystic fibrosis transmembrane conductance regulator. / Akabas, Myles.

In: Biochemistry, Vol. 37, No. 35, 01.09.1998, p. 12233-12240.

Research output: Contribution to journalArticle

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