title: Biophysical effects of pore mutations of ROMK1.
abstract: Potassium channels are ubiquitous, being present in all living organisms. These proteins share common structural elements, which confer common functional features. In general, all K+ channels have a high selectivity for K+, and are blocked by cations of similar dimensions, such as Cs+ and Ba2+. Mutations in the pore region tend to lead to either the total loss of function or K+ selectivity. We have made mutations to one of the most highly conserved residues of the pore, glycine-143, of the inward rectifier ROMK1 (Kir1.1), and examined the resulting channel properties in the Xenopus oocyte expression system with a two-electrode voltage clamp. Mutations G143A and G143R resulted in failure to express functional channels. Co-injection of wild-type ROMK1 cRNA with these mutants led to rescue of channel function, which was different from wild-type ROMK1. In both mutants, the sensitivity to Ba2+ and Cs+ was increased, the rate of onset of block by Ba2+ was enhanced, and the selectivity to potassium was reduced. Whereas the crystallographic evidence shows that cations bind to the carbonyl backbone of the pore-lining residues, the present results indicate that the side chains of these amino acids, which face away from the pore lining, also affect permeation.
authors.affiliation: School of Biomedical Sciences, Department of Physiology, Worsley Medical Building, University of Leeds, Leeds LS2 9NQ, UK. sunil.bhandari@medix-uk.com
authors.forename: S
authors.initials: S
authors.lastname: Bhandari
authors.forename: M
authors.initials: M
authors.lastname: Hunter
mesh_terms: D000818:Animals; D001464:Barium / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000235:genetics; D002586:Cesium / Q000378:metabolism; D005260:Female; D005998:Glycine / Q000235:genetics*; D016014:Linear Models; D009154:Mutation / Q000235:genetics*; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D016133:Polymerase Chain Reaction / Q000379:methods; D015221:Potassium Channels / Q000235:genetics* / Q000502:physiology; D024661:Potassium Channels, Inwardly Rectifying; D018075:RNA, Complementary / Q000235:genetics; D014981:Xenopus
chemical_list: D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D018075:RNA, Complementary; D002586:Cesium; D001464:Barium; D005998:Glycine