title: Interactions of cations with the cytoplasmic pores of inward rectifier K(+) channels in the closed state.
abstract: Ion channels gate at membrane-embedded domains by changing their conformation along the ion conduction pathway. Inward rectifier K(+) (Kir) channels possess a unique extramembrane cytoplasmic domain that extends this pathway. However, the relevance and contribution of this domain to ion permeation remain unclear. By qualitative x-ray crystallographic analysis, we found that the pore in the cytoplasmic domain of Kir3.2 binds cations in a valency-dependent manner and does not allow the displacement of Mg(2+) by monovalent cations or spermine. Electrophysiological analyses revealed that the cytoplasmic pore of Kir3.2 selectively binds positively charged molecules and has a higher affinity for Mg(2+) when it has a low probability of being open. The selective blocking of chemical modification of the side chain of pore-facing residues by Mg(2+) indicates that the mode of binding of Mg(2+) is likely to be similar to that observed in the crystal structure. These results indicate that the Kir3.2 crystal structure has a closed conformation with a negative electrostatic field potential at the cytoplasmic pore, the potential of which may be controlled by conformational changes in the cytoplasmic domain to regulate ion diffusion along the pore.
authors: Department of Pharmacology, Graduate School of Medicine, Osaka University, Osaka 565-0871, Japan; Center for Advanced Medical Engineering and Informatics, Osaka University, Osaka 565-0871, Japan. Electronic address: inanobe@pharma2.med.osaka-u.ac.jp.
authors: Atsushi
authors: A
authors: Inanobe
authors: Laboratory of Supramolecular Crystallography, Institute for Protein Research, Osaka University, Osaka 565-0871, Japan.
authors: Atsushi
authors: A
authors: Nakagawa
authors: Department of Pharmacology, Graduate School of Medicine, Osaka University, Osaka 565-0871, Japan; Center for Advanced Medical Engineering and Informatics, Osaka University, Osaka 565-0871, Japan. Electronic address: ykurachi@pharma2.med.osaka-u.ac.jp.
authors: Yoshihisa
authors: Y
authors: Kurachi
mesh_terms: D000818:Animals; D002413:Cations, Divalent / Q000737:chemistry* / Q000378:metabolism; D018360:Crystallography, X-Ray; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008274:Magnesium / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D017434:Protein Structure, Tertiary
chemical_list: D002413:Cations, Divalent; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; C497069:Kcnj6 protein, mouse; D008274:Magnesium