Structure of the BK potassium channel in a lipid membrane from electron cryomicroscopy

• Published in: Nature (London) Vol. 461; no. 7261; pp. 292 - 295
• Main Authors: LIGUO WANG, SIGWORTH, Fred J
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group 10.09.2009
• Subjects: Atoms & subatomic particles; Biological and medical sciences; Cell Line; Chemical properties; Cryoelectron Microscopy - methods; Data collection; Electrons; Fluorescence; Fundamental and applied biological sciences. Psychology; Humans; Ion Channel Gating; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits - chemistry; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits - genetics; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits - metabolism; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits - ultrastructure; Lipid membranes; Lipids; Liposomes - chemistry; Liposomes - metabolism; Membrane Lipids - metabolism; Membrane Potentials; Membranes; Models, Molecular; Molecular biophysics; Observations; Potassium; Potassium - metabolism; Potassium channels; Protein Structure, Tertiary; Proteins; Proteolipids - chemistry; Proteolipids - metabolism; Structure in molecular biology; Substrate Specificity; Tridimensional structure; United States; Membrane protein; Voltage-gated canal; Three dimensional structure; Liposome; cryo electron microscopy; Membrane; Ionic channel; Potassium; Protein
• ISSN: 0028-0836; 1476-4687
• DOI: 10.1038/nature08291

A long-sought goal in structural biology has been the imaging of membrane proteins in their membrane environments. This goal has been achieved with electron crystallography in those special cases where a protein forms highly ordered arrays in lipid bilayers. It has also been achieved by NMR methods in proteins up to 50 kilodaltons (kDa) in size, although milligram quantities of protein and isotopic labelling are required. For structural analysis of large soluble proteins in microgram quantities, an increasingly powerful method that does not require crystallization is single-particle reconstruction from electron microscopy of cryogenically cooled samples (electron cryomicroscopy (cryo-EM)). Here we report the first single-particle cryo-EM study of a membrane protein, the human large-conductance calcium- and voltage-activated potassium channel (BK), in a lipid environment. The new method is called random spherically constrained (RSC) single-particle reconstruction. BK channels, members of the six-transmembrane-segment (6TM) ion channel family, were reconstituted at low density into lipid vesicles (liposomes), and their function was verified by a potassium flux assay. Vesicles were also frozen in vitreous ice and imaged in an electron microscope. From images of 8,400 individual protein particles, a three-dimensional (3D) reconstruction of the BK channel and its membrane environment was obtained at a resolution of 1.7-2.0 nm. Not requiring the formation of crystals, the RSC approach promises to be useful in the structural study of many other membrane proteins as well.