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The selectivity filter of a potassium channel, murine Kir2.1, investigated using scanning cysteine mutagenesis
Author(s) -
Dart C.,
Leyland M. L.,
Spencer P. J.,
Stanfield P. R.,
Sutcliffe M. J.
Publication year - 1998
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1111/j.1469-7793.1998.025bi.x
Subject(s) - tetramer , cysteine , mutagenesis , chemistry , potassium channel , mutant , covalent bond , side chain , inward rectifier potassium ion channel , biophysics , selectivity , ion channel , kcsa potassium channel , stereochemistry , biochemistry , biology , gene , organic chemistry , receptor , catalysis , enzyme , polymer
1 We have produced a structural model of the pore‐forming H5 (or P) region of the strong inward rectifier K + channel, Kir2.1, based initially on an existing molecular model of the pore region of the voltage‐gated K + channel, Kv1.3. 2 Cysteine‐scanning mutagenesis and subsequent blockage by Ag + was used to test our model by determining the residues in H5 whose side chains line the ion conduction pathway. 3 Mutations made in eight positions within the highly conserved H5 region resulted in apparently non‐functional channels. Constructing covalently linked dimers, which carry a cysteine substitution in only one of the linked subunits, rescued six of these mutants; a covalently linked tetramer, carrying a cysteine substitution on only one of the linked subunits, rescued a further mutant. 4 Our results using the dimers and tetramers suggest that residues Thr141, Thr142, Ile143, Tyr145, Phe147 and Cys149 are accessible to externally applied Ag + (100‐200 nM) and therefore that their side chains line the channel pore. 5 We conclude that the topology of the Kir pore is similar, but not identical, to that of Kv channels. Additionally, the molecular model suggests that selectivity may be conferred both by aromatic residues (Tyr145 and Phe147) via cation‐π interactions and by backbone carbonyl groups (Thr142 and Gly144).