title: Intracellular segment between transmembrane helices S0 and S1 of BK channel α subunit contains two amphipathic helices connected by a flexible loop.
abstract: The BK channel, a tetrameric potassium channel with very high conductance, has a central role in numerous physiological functions. The BK channel can be activated by intracellular Ca(2+) and Mg(2+), as well as by membrane depolarization. Unlike other tetrameric potassium channels, the BK channel has seven transmembrane helices (S0-S6) including an extra helix S0. The intracellular segment between S0 and S1 (BK-IS1) is essential to BK channel functions and Asp99 in BK-IS1 is reported to be responsible for Mg(2+) coordination. In this study, BK-IS1 (44-113) was over-expressed using a bacterial system and purified in the presence of detergent micelles for multidimensional heteronuclear nuclear magnetic resonance (NMR) structural studies. Backbone resonance assignment and secondary structure analysis showed that BK-IS1 contains two amphipathic helices connected by a 36-residue loop. Amide (1)H-(15)N heteronuclear NOE analysis indicated that the loop is very flexible, while the two amphipathic helices are possibly stabilized through interaction with the membrane. A solution NMR-based titration assay of BK-IS1 was performed with various concentrations of Mg(2+). Two residues (Thr45 and Leu46) with chemical shift changes were observed but no, or very minor, chemical shift difference was observed for Asp99, indicating a possible site for binding divalent ions or other modulation partners.
authors.affiliation: Hefei National Laboratory of Microscale Physical Sciences, School of Life Sciences, University of Science and Technology of China, Hefei, Anhui, 230027, PR China.
authors.forename: Pan
authors.initials: P
authors.lastname: Shi
authors.forename: Dong
authors.initials: D
authors.lastname: Li
authors.forename: Chaohua
authors.initials: C
authors.lastname: Lai
authors.forename: Longhua
authors.initials: L
authors.lastname: Zhang
authors.forename: Changlin
authors.initials: C
authors.lastname: Tian
mesh_terms: D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D006801:Humans; D007425:Intracellular Membranes / Q000737:chemistry*; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits / Q000737:chemistry* / Q000235:genetics; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000737:chemistry* / Q000235:genetics; D008051:Lipid Bilayers / Q000737:chemistry; D008823:Micelles; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics; D010713:Phosphatidylcholines / Q000737:chemistry; D010767:Phosphorylcholine / Q000031:analogs & derivatives / Q000737:chemistry; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics
chemical_list: D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D051036:Large-Conductance Calcium-Activated Potassium Channels; D008051:Lipid Bilayers; D008823:Micelles; D010446:Peptide Fragments; D010713:Phosphatidylcholines; D021122:Protein Subunits; D010767:Phosphorylcholine; C028810:dodecylphosphocholine
keywords: BK channel; Backbone assignment; Intracellular segment; Secondary structure; Solution NMR; Titration