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.
???displayArticle.abstract???
We report the use of cysteine-substituted mutants in conjunction with in situ oxidation to determine the physical proximity of a pair of engineered cysteines in the pore region of the voltage-gated K+ channel Kv2.1. We show that the newly introduced cysteine 1379C, located near the outer end of the narrow ion-conduction pathway, renders the K+ channel sensitive to oxidation by H2O2, but only if the native cysteine at position 394 in S6 remains in place. Conservative substitutions in S6 for cysteine 394 abolish H2O2 sensitivity in the Kv2.1 mutant 1379C. Comparative immunoblot analysis of wild-type and 1379C Kv2.1-expressing HEK293 cells demonstrates the presence of subunit dimers for 1379C, but not for wild-type Kv2.1. At the single-channel level, the probability of opening of 1379C channels, unlike wild-type, is reduced in the presence of H2O2; however, oxidation of 1379C does not alter unit current. These findings imply that cysteine 379, located near the outer end of the narrow ion-conduction pathway, participates in disulfide bridge formation, locking the channel in a nonconducting state from which it cannot undergo conformational transitions required for opening.
Aiyar,
Topology of the pore-region of a K+ channel revealed by the NMR-derived structures of scorpion toxins.
1995, Pubmed
Aiyar,
Topology of the pore-region of a K+ channel revealed by the NMR-derived structures of scorpion toxins.
1995,
Pubmed
Bezanilla,
Molecular basis of gating charge immobilization in Shaker potassium channels.
1991,
Pubmed
,
Xenbase
Careaga,
Structure and dynamics of Escherichia coli chemosensory receptors. Engineered sulfhydryl studies.
1992,
Pubmed
Choi,
The internal quaternary ammonium receptor site of Shaker potassium channels.
1993,
Pubmed
,
Xenbase
Frech,
A novel potassium channel with delayed rectifier properties isolated from rat brain by expression cloning.
1989,
Pubmed
,
Xenbase
Hartmann,
Exchange of conduction pathways between two related K+ channels.
1991,
Pubmed
,
Xenbase
Hoshi,
Two types of inactivation in Shaker K+ channels: effects of alterations in the carboxy-terminal region.
1991,
Pubmed
,
Xenbase
Kavanaugh,
Interaction between tetraethylammonium and amino acid residues in the pore of cloned voltage-dependent potassium channels.
1991,
Pubmed
,
Xenbase
Kürz,
Side-chain accessibilities in the pore of a K+ channel probed by sulfhydryl-specific reagents after cysteine-scanning mutagenesis.
1995,
Pubmed
,
Xenbase
Liman,
Subunit stoichiometry of a mammalian K+ channel determined by construction of multimeric cDNAs.
1992,
Pubmed
,
Xenbase
Liu,
Dynamic rearrangement of the outer mouth of a K+ channel during gating.
1996,
Pubmed
Lopez,
Evidence that the S6 segment of the Shaker voltage-gated K+ channel comprises part of the pore.
1994,
Pubmed
,
Xenbase
Lü,
Silver as a probe of pore-forming residues in a potassium channel.
1995,
Pubmed
,
Xenbase
Lynch,
Disulfide cross-linking studies of the transmembrane regions of the aspartate sensory receptor of Escherichia coli.
1991,
Pubmed
MacKinnon,
Determination of the subunit stoichiometry of a voltage-activated potassium channel.
1991,
Pubmed
,
Xenbase
Moorman,
Fast and slow gating of sodium channels encoded by a single mRNA.
1990,
Pubmed
,
Xenbase
Pakula,
Determination of transmembrane protein structure by disulfide cross-linking: the Escherichia coli Tar receptor.
1992,
Pubmed
Pascual,
K+ pore structure revealed by reporter cysteines at inner and outer surfaces.
1995,
Pubmed
,
Xenbase
Rudy,
Diversity and ubiquity of K channels.
1988,
Pubmed
Slesinger,
The S4-S5 loop contributes to the ion-selective pore of potassium channels.
1993,
Pubmed
,
Xenbase
VanDongen,
Alteration and restoration of K+ channel function by deletions at the N- and C-termini.
1990,
Pubmed
Yellen,
An engineered cysteine in the external mouth of a K+ channel allows inactivation to be modulated by metal binding.
1994,
Pubmed
Yellen,
Mutations affecting internal TEA blockade identify the probable pore-forming region of a K+ channel.
1991,
Pubmed
Yool,
Alteration of ionic selectivity of a K+ channel by mutation of the H5 region.
1991,
Pubmed
,
Xenbase
Zühlke,
Role of an invariant cysteine in gating and ion permeation of the voltage-sensitive K+ channel Kv2.1.
1994,
Pubmed
,
Xenbase
