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Electrophysiological, Biochemical, and Bioinformatic Methods for Studying CFTR Channel Gating and Its Regulation

  • László CsanádyEmail author
  • Paola Vergani
  • Attila Gulyás-Kovács
  • David C. GadsbyEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 741)

Abstract

CFTR is the only member of the ABC (ATP-binding cassette) protein superfamily known to function as an ion channel. Most other ABC proteins are ATP-driven transporters, in which a cycle of ATP binding and hydrolysis, at intracellular nucleotide binding domains (NBDs), powers uphill substrate translocation across the membrane. In CFTR, this same ATP-driven cycle opens and closes a transmembrane pore through which chloride ions flow rapidly down their electrochemical gradient. Detailed analysis of the pattern of gating of CFTR channels thus offers the opportunity to learn about mechanisms of function not only of CFTR channels but also of their ABC transporter ancestors. In addition, CFTR channel gating is subject to complex regulation by kinase-mediated phosphorylation at multiple consensus sites in a cytoplasmic regulatory domain that is unique to CFTR. Here we offer a practical guide to extract useful information about the mechanisms that control opening and closing of CFTR channels: on how to plan (including information obtained from analysis of multiple sequence alignments), carry out, and analyze electrophysiological and biochemical experiments, as well as on how to circumvent potential pitfalls.

Key words

Single channels multiple channels kinetic analysis energetic analysis cysteine modification coevolution 

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  1. 1.Department of Medical BiochemistrySemmelweis UniversityBudapestHungary
  2. 2.Department of Neuroscience, Physiology and PharmacologyUniversity College LondonLondonUK
  3. 3.Laboratory of Cardiac/Membrane PhysiologyThe Rockefeller UniversityNew YorkUSA
  4. 4.Laboratory of Cardiac/Membrane PhysiologyThe Rockefeller UniversityNew YorkUSA

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