title: Crystal structure of a Ba(2+)-bound gating ring reveals elementary steps in RCK domain activation.
abstract: RCK domains control activity of a variety of K(+) channels and transporters through binding of cytoplasmic ligands. To gain insight toward mechanisms of RCK domain activation, we solved the structure of the RCK domain from the Ca(2+)-gated K(+) channel, MthK, bound with Ba(2+), at 3.1 Å resolution. The Ba(2+)-bound RCK domain was assembled as an octameric gating ring, as observed in structures of the full-length MthK channel, and shows Ba(2+) bound at several positions. One of the Ba(2+) sites, termed C1, overlaps with a known Ca(2+)-activation site, determined by residues D184 and E210. Functionally, Ba(2+) can activate reconstituted MthK channels as observed in electrophysiological recordings, whereas Mg(2+) (up to 100 mM) was ineffective. Ba(2+) activation was abolished by the mutation D184N, suggesting that Ba(2+) activates primarily through the C1 site. Our results suggest a working hypothesis for a sequence of ligand-dependent conformational changes that may underlie RCK domain activation and channel gating.
authors.affiliation: Department of Biochemistry, Temple University School of Medicine, 3400 N. Broad Street, Philadelphia, PA 19140, USA.
authors.forename: Frank J
authors.initials: FJ
authors.lastname: Smith
authors.forename: Victor P T
authors.initials: VP
authors.lastname: Pau
authors.forename: Gino
authors.initials: G
authors.lastname: Cingolani
authors.forename: Brad S
authors.initials: BS
authors.lastname: Rothberg
mesh_terms: D020816:Amino Acid Motifs; D019843:Archaeal Proteins / Q000737:chemistry*; D001464:Barium / Q000737:chemistry*; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry; D056831:Coordination Complexes / Q000737:chemistry; D018360:Crystallography, X-Ray; D006863:Hydrogen-Ion Concentration; D015640:Ion Channel Gating; D008051:Lipid Bilayers / Q000737:chemistry; D008564:Membrane Potentials; D017013:Methanobacteriaceae; D008958:Models, Molecular; D010714:Phosphatidylethanolamines / Q000737:chemistry; D010715:Phosphatidylglycerols / Q000737:chemistry; D024681:Potassium Channels, Calcium-Activated / Q000737:chemistry*; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary
chemical_list: D019843:Archaeal Proteins; D056831:Coordination Complexes; D008051:Lipid Bilayers; D010714:Phosphatidylethanolamines; D010715:Phosphatidylglycerols; D024681:Potassium Channels, Calcium-Activated; C057561:1-palmitoyl-2-oleoylphosphatidylethanolamine; D001464:Barium; C060037:1-palmitoyl-2-oleoylglycero-3-phosphoglycerol; D002118:Calcium