CFTR (Cystic Fibrosis Transmembrane conductance Regulator) is a chloride channel with unique properties. Being a member of the ABC (ATP Binding Cassette) transporter superfamily, CFTR uses ATP hydrolysis as the energy source to carry out its function. Unlike other members of this family that use the free energy of ATP hydrolysis to transport substrates against the electrochemical gradients, CFTR harvests the free energy to drive its conformational changes during opening and closing (or gating) of the channel. Proteins in this family play numerous physiological and pathophysiological roles in a variety of systems including epithelial chloride secretion, transport of cholesterol, drug resistance in cancers, cardiac membrane excitability and insulin secretion. Thus, understanding how CFTR works at a molecular level will have a broad impact on both basic sciences and clinical medicine. ? ? Recent solution of X-ray crystal structure of CFTR's N-terminal nucleotide binding domain (NBD1) has opened the door for detailed studies of the role of NBDs in controlling gating transitions. The current proposal will employ a combination of electrophysiological, molecular biological, and structural biological techniques to address some fundamental questions of CFTR gating: What is the role of individual NBDs in modulating CFTR gating? What is the chemical nature of interactions between ATP and its binding pockets? Is binding of ATP at both NBDs absolutely required for channel opening? Since NBD1 lacks the essential amino acids for ATP hydrolysis, what is the role of ATP binding at NBD1? Our specific aims are:
Aim 1. To study CFTR gating kinetics using structure-guided mutagenesis.
Aim 2. To determine the kinetic and energetic roles of individual NBDs in CFTR gating using novel nucleotide analogs. A clear understanding of the molecular mechanisms of CFTR function will aid in designs of therapeutical reagents for the treatment of cystic fibrosis, secretory diarrhea, and other CFTR-associated diseases. ? ? ?
|Sohma, Yoshiro; Yu, Ying-Chun; Hwang, Tzyh-Chang (2013) Curcumin and genistein: the combined effects on disease-associated CFTR mutants and their clinical implications. Curr Pharm Des 19:3521-8|
|Hwang, Tzyh-Chang; Kirk, Kevin L (2013) The CFTR ion channel: gating, regulation, and anion permeation. Cold Spring Harb Perspect Med 3:a009498|
|Jih, Kang-Yang; Hwang, Tzyh-Chang (2012) Nonequilibrium gating of CFTR on an equilibrium theme. Physiology (Bethesda) 27:351-61|
|Jih, Kang-Yang; Sohma, Yoshiro; Li, Min et al. (2012) Identification of a novel post-hydrolytic state in CFTR gating. J Gen Physiol 139:359-70|
|Bai, Yonghong; Li, Min; Hwang, Tzyh-Chang (2011) Structural basis for the channel function of a degraded ABC transporter, CFTR (ABCC7). J Gen Physiol 138:495-507|
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|Tsai, Ming-Feng; Jih, Kang-Yang; Shimizu, Hiroyasu et al. (2010) Optimization of the degenerated interfacial ATP binding site improves the function of disease-related mutant cystic fibrosis transmembrane conductance regulator (CFTR) channels. J Biol Chem 285:37663-71|
|Tsai, Ming-Feng; Li, Min; Hwang, Tzyh-Chang (2010) Stable ATP binding mediated by a partial NBD dimer of the CFTR chloride channel. J Gen Physiol 135:399-414|
|Shimizu, Hiroyasu; Yu, Ying-Chun; Kono, Koichi et al. (2010) A stable ATP binding to the nucleotide binding domain is important for reliable gating cycle in an ABC transporter CFTR. J Physiol Sci 60:353-62|
|Hwang, Tzyh-Chang; Sheppard, David N (2009) Gating of the CFTR Cl- channel by ATP-driven nucleotide-binding domain dimerisation. J Physiol 587:2151-61|
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