Mutations at the GABA receptor selectivity filter: A possible role for effective charges

V. E. Wotring, T. S. Miller, D. S. Weiss

Research output: Contribution to journalArticlepeer-review

47 Scopus citations

Abstract

An important feature of ligand-gated ion channels is their exquisite ability to discriminate between ions. Still, little is known about the mechanisms underlying, or structural determinates of, this ability. We examined the structural elements underlying the ionic selectivity of ρ1 GABA receptors expressed in Xenopus oocytes and human embryonic kidney cells using site-directed mutagenesis and two-electrode voltage-clamp or patch-clamp techniques. The wild-type GABA receptor was chloride selective, with a small but significant permeability to potassium of the second transmembrane domain), formed a channel that exhibited little discrimination among ions (0.70:0.87:1), while deletion of a neighbouring proline (290) was chloride selective, but had elevated cation permeabilities compared to the wild-type channel (0.12:0.14:1). Together, the two mutations (ΔP290/A291E) caused a reversal of selectivity (2.72:3.59:1). We also examined the effects of neutralizing and reversing the charge of the adjacent, and highly conserved, arginine. Mutation of the neighbouring arginine to glutamate (R292E) increased the cation permeability similar to the ΔP290/A291E double mutant (2.4:3.0:1), whereas neutral mutations at this position (R292M or R292C) retained chloride selectivity (0:0.11:1.0 and 0:0.14:1.0, respectively). Our experiments suggest that the effective charge near the presumed intracellular mouth of the pore is critical for ionic selectivity.

Original languageEnglish (US)
Pages (from-to)527-540
Number of pages14
JournalJournal of Physiology
Volume548
Issue number2
DOIs
StatePublished - Apr 15 2003

ASJC Scopus subject areas

  • Physiology

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