title: Metal-driven operation of the human large-conductance voltage- and Ca2+-dependent potassium channel (BK) gating ring apparatus.
abstract: Large-conductance voltage- and Ca(2+)-dependent K(+) (BK, also known as MaxiK) channels are homo-tetrameric proteins with a broad expression pattern that potently regulate cellular excitability and Ca(2+) homeostasis. Their activation results from the complex synergy between the transmembrane voltage sensors and a large (>300 kDa) C-terminal, cytoplasmic complex (the "gating ring"), which confers sensitivity to intracellular Ca(2+) and other ligands. However, the molecular and biophysical operation of the gating ring remains unclear. We have used spectroscopic and particle-scale optical approaches to probe the metal-sensing properties of the human BK gating ring under physiologically relevant conditions. This functional molecular sensor undergoes Ca(2+)- and Mg(2+)-dependent conformational changes at physiologically relevant concentrations, detected by time-resolved and steady-state fluorescence spectroscopy. The lack of detectable Ba(2+)-evoked structural changes defined the metal selectivity of the gating ring. Neutralization of a high-affinity Ca(2+)-binding site (the "calcium bowl") reduced the Ca(2+) and abolished the Mg(2+) dependence of structural rearrangements. In congruence with electrophysiological investigations, these findings provide biochemical evidence that the gating ring possesses an additional high-affinity Ca(2+)-binding site and that Mg(2+) can bind to the calcium bowl with less affinity than Ca(2+). Dynamic light scattering analysis revealed a reversible Ca(2+)-dependent decrease of the hydrodynamic radius of the gating ring, consistent with a more compact overall shape. These structural changes, resolved under physiologically relevant conditions, likely represent the molecular transitions that initiate the ligand-induced activation of the human BK channel.
authors.affiliation: Division of Molecular Medicine, Department of Anesthesiology, David Geffen School of Medicine, UCLA, Los Angeles, California 90095-7115, USA.
authors.forename: Anoosh D
authors.initials: AD
authors.lastname: Javaherian
authors.forename: Taleh
authors.initials: T
authors.lastname: Yusifov
authors.forename: Antonios
authors.initials: A
authors.lastname: Pantazis
authors.forename: Sarah
authors.initials: S
authors.lastname: Franklin
authors.forename: Chris S
authors.initials: CS
authors.lastname: Gandhi
authors.forename: Riccardo
authors.initials: R
authors.lastname: Olcese
mesh_terms: D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D015640:Ion Channel Gating / Q000502:physiology*; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008274:Magnesium / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence
chemical_list: D051036:Large-Conductance Calcium-Activated Potassium Channels; D008274:Magnesium; D002118:Calcium