Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Biochemistry
2011 Nov 29;5047:10311-7. doi: 10.1021/bi201176q.
Show Gene links
Show Anatomy links
Cystic fibrosis transmembrane conductance regulator: temperature-dependent cysteine reactivity suggests different stable conformers of the conduction pathway.
Liu X
,
Dawson DC
.
???displayArticle.abstract???
Cysteine scanning has been widely used to identify pore-lining residues in mammalian ion channels, including the cystic fibrosis transmembrane conductance regulator (CFTR). These studies, however, have been typically conducted at room temperature rather than human body temperature. Reports of substantial effects of temperature on gating and anion conduction in CFTR channels as well as an unexpected pattern of cysteine reactivity in the sixth transmembrane segment (TM6) prompted us to investigate the effect of temperature on the reactivity of cysteines engineered into TM6 of CFTR. We compared reaction rates at temperatures ranging from 22 to 37 °C for cysteines placed on either side of an apparent size-selective accessibility barrier previously defined by comparing reactivity toward channel-permeant and channel-impermeant, thiol-directed reagents. The results indicate that the reactivity of cysteines at three positions extracellular to the position of the accessibility barrier, 334, 336, and 337, is highly temperature-dependent. At 37 °C, cysteines at these positions were highly reactive toward MTSES(-), whereas at 22 °C, the reaction rates were 2-6-fold slower to undetectable. An activation energy of 157 kJ/mol for the reaction at position 337 is consistent with the hypothesis that, at physiological temperature, the extracellular portion of the CFTR pore can adopt conformations that differ significantly from those that can be accessed at room temperature. However, the position of the accessibility barrier defined empirically by applying channel-permeant and channel-impermeant reagents to the extracellular aspect of the pore is not altered. The results illuminate previous scanning results and indicate that the assay temperature is a critical variable in studies designed to use chemical modification to test structural models for the CFTR anion conduction pathway.
Alexander,
Cystic fibrosis transmembrane conductance regulator: using differential reactivity toward channel-permeant and channel-impermeant thiol-reactive probes to test a molecular model for the pore.
2009, Pubmed,
Xenbase
Alexander,
Cystic fibrosis transmembrane conductance regulator: using differential reactivity toward channel-permeant and channel-impermeant thiol-reactive probes to test a molecular model for the pore.
2009,
Pubmed
,
Xenbase
Bai,
Dual roles of the sixth transmembrane segment of the CFTR chloride channel in gating and permeation.
2010,
Pubmed
Beck,
Conformational changes in a pore-lining helix coupled to cystic fibrosis transmembrane conductance regulator channel gating.
2008,
Pubmed
,
Xenbase
Bischof,
Thermal stability of proteins.
2005,
Pubmed
Bodnar,
Transition state characterization of the low- to physiological-temperature nondenaturational conformational change in bovine adenosine deaminase by slow scan rate differential scanning calorimetry.
2006,
Pubmed
Britto,
The local electrostatic environment determines cysteine reactivity of tubulin.
2002,
Pubmed
Cheng,
Defective intracellular transport and processing of CFTR is the molecular basis of most cystic fibrosis.
1990,
Pubmed
Cheung,
Identification of cystic fibrosis transmembrane conductance regulator channel-lining residues in and flanking the M6 membrane-spanning segment.
1996,
Pubmed
,
Xenbase
Cheung,
Locating the anion-selectivity filter of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel.
1997,
Pubmed
Csanády,
Thermodynamics of CFTR channel gating: a spreading conformational change initiates an irreversible gating cycle.
2006,
Pubmed
,
Xenbase
El Hiani,
Changes in accessibility of cytoplasmic substances to the pore associated with activation of the cystic fibrosis transmembrane conductance regulator chloride channel.
2010,
Pubmed
Fatehi,
State-dependent access of anions to the cystic fibrosis transmembrane conductance regulator chloride channel pore.
2008,
Pubmed
Gentzsch,
Endocytic trafficking routes of wild type and DeltaF508 cystic fibrosis transmembrane conductance regulator.
2004,
Pubmed
Heda,
The Delta F508 mutation shortens the biochemical half-life of plasma membrane CFTR in polarized epithelial cells.
2001,
Pubmed
Hollowell,
Thermodynamic analysis of the low- to physiological-temperature nondenaturational conformational change of bovine carbonic anhydrase.
2007,
Pubmed
Iversen,
Thiol-disulfide exchange between glutaredoxin and glutathione.
2010,
Pubmed
Karlin,
Substituted-cysteine accessibility method.
1998,
Pubmed
Liang,
Kinetic measurements on single-molecule disulfide bond cleavage.
2011,
Pubmed
Liu,
Variable reactivity of an engineered cysteine at position 338 in cystic fibrosis transmembrane conductance regulator reflects different chemical states of the thiol.
2006,
Pubmed
,
Xenbase
Liu,
CFTR: a cysteine at position 338 in TM6 senses a positive electrostatic potential in the pore.
2004,
Pubmed
,
Xenbase
Ma,
Thiazolidinone CFTR inhibitor identified by high-throughput screening blocks cholera toxin-induced intestinal fluid secretion.
2002,
Pubmed
Massey,
A temperature-dependent conformational change in D-amino acid oxidase and its effect on catalysis.
1966,
Pubmed
Mathews,
The CFTR chloride channel: nucleotide interactions and temperature-dependent gating.
1998,
Pubmed
Okiyoneda,
Peripheral protein quality control removes unfolded CFTR from the plasma membrane.
2010,
Pubmed
Serrano,
CFTR: Ligand exchange between a permeant anion ([Au(CN)2]-) and an engineered cysteine (T338C) blocks the pore.
2006,
Pubmed
,
Xenbase
Sharma,
Misfolding diverts CFTR from recycling to degradation: quality control at early endosomes.
2004,
Pubmed
Sharma,
Conformational and temperature-sensitive stability defects of the delta F508 cystic fibrosis transmembrane conductance regulator in post-endoplasmic reticulum compartments.
2001,
Pubmed
Smith,
CFTR: covalent and noncovalent modification suggests a role for fixed charges in anion conduction.
2001,
Pubmed
,
Xenbase
Snyder,
Electrostatic influence of local cysteine environments on disulfide exchange kinetics.
1981,
Pubmed
Varga,
Enhanced cell-surface stability of rescued DeltaF508 cystic fibrosis transmembrane conductance regulator (CFTR) by pharmacological chaperones.
2008,
Pubmed
Wang,
Alignment of transmembrane regions in the cystic fibrosis transmembrane conductance regulator chloride channel pore.
2011,
Pubmed
