Structure and insights into the function of a Ca2+-activated Cl− channel

Dickson, Veronica Kane; Pedi, Leanne; Long, Stephen B.

doi:10.1038/nature13913

Cited by 185 publications

(312 citation statements)

References 61 publications

Supporting

Mentioning

299

Contrasting

Unclassified

Order By: Relevance

“…One example is the pentameric bestrophin channel in which three layers of hydrophobic residues create a 10Å long, 3Å wide ion pathway resembling that of CmTMEM175 (Extended Data Fig. 5) 22,23 . Intriguingly, the animal Bestrophin channel, whose narrow pathway is formed by three layers of Ile, Phe, and Phe, is an anion selective channel while the bacterial Bestrophin, which contains Ile, Ile, and Phe at the bottle neck, has been shown to be cation selective channel 22,24,25 .…”

mentioning

confidence: 99%

The lysosomal potassium channel TMEM175 adopts a novel tetrameric architecture

Lee

Guo

Zeng

et al. 2017

Nature

View full text Add to dashboard Cite

TMEM175 is a lysosomal K+ channel important for maintaining the lysosomal membrane potential and pH stability1. It contains two homologous copies of a six-transmembrane (6-TM) domain, which has no sequence homology to the canonical tetrameric K+ channels and lacks the TVGYG selectivity filter motif2–4. Present in a subset of bacteria and archaea, the prokaryotic TMEM175 contains only a single 6-TM domain and functions as a tetramer. Here, we present the crystal structure of a prokaryotic TMEM175 from Chamaesiphon minutus, CmTMEM175, whose architecture represents a completely different fold from that of canonical K+ channels. All six transmembrane helices of CmTMEM175 are tightly packed within each subunit without undergoing domain swap. The highly conserved TM1 acts as the pore lining inner helix, creating an hour-glass shaped ion permeation pathway in the channel tetramer. Three layers of hydrophobic residues on the C-terminal half of TM1s form a bottle neck along the ion conduction pathway and serve as the selectivity filter of the channel. Mutagenesis analysis suggests that the first layer of the highly conserved isoleucine residues in the filter plays the central role in channel selectivity. Thus, the structure of CmTMEM175 represents a novel architecture of a tetrameric cation channel whose ion selectivity mechanism appears distinct from that of the classical K+ channel family.

show abstract

mentioning

confidence: 99%

The lysosomal potassium channel TMEM175 adopts a novel tetrameric architecture

Lee

Guo

Zeng

et al. 2017

Nature

View full text Add to dashboard Cite

show abstract

“…This residue was selected in part owing to a report that calcium binding suffered if the protein was shorter than residues 1-380 (human Best1 numbering; Xiao et al, 2008), and the sequence conservation was increasingly weak beyond residue 398. Protein that was truncated at this point was shown to be active using an assay of purified, reconstituted protein (Kane Dickson et al, 2014). Of the 30 eukaryotic candidate orthologues expressed as a GFP fusion protein in HEK293 cells and screened by fluorescence sizeexclusion chromatography (FSEC; Kawate & Gouaux, 2006), three were selected for further testing.…”

Section: Candidate Selectionmentioning

confidence: 99%

“…The purified channel demonstrated good characteristics as assessed by gel filtration and was functional as demonstrated by a flux assay (Kane Dickson et al, 2014). A thermostability assay was carried out in the presence of a range of specific inhibitors and ions using analytical size-exclusion chromatography (SEC).…”

Section: Purification and Initial Crystallizationmentioning

confidence: 99%

“…The protein was solubilized in n-dodecyl--d-maltopyranoside (DDM) and purified in DDM or another maltoside for crystallization (Kane Dickson et al, 2014). Similar to the GFP-labelled protein in FSEC screening, the protein purified from Pichia also demonstrated stability over a range of detergents and formed crystals in several detergents tested.…”

Section: Purification and Initial Crystallizationmentioning

confidence: 99%

“…Optimized crystals which contained the GgBest1-Fab complex were grown by vapour diffusion either in CYMAL-6 or CYMAL-6 neopentyl glycol (CYMAL-6-NG) against a well solution consisting of 0-60 mM NaCl, 50 mM sodium acetate pH 4.0, 5% PEG 4000, 20% glycerol, generating crystals of the P2 1 crystal form, or in DM against a well solution consisting of 120 mM NaCl, 50 mM Tris pH 8.5, 8.5% PEG 4000, 20% glycerol, generating crystals of the C2 form (Kane Dickson et al, 2014). Both forms were grown by hanging-drop vapour diffusion using a 1:1 ratio of protein solution to well solution either in 100-600 nl drops (96-well format) or in 0.8-3.8 ml drops (24-well format).…”

Section: Crystallization and Heavy-atom Derivatization Trialsmentioning

confidence: 99%

See 2 more Smart Citations

Phasing and structure of bestrophin-1: a case study in the use of heavy-atom cluster compounds with multi-subunit transmembrane proteins

Dickson

2016

Acta Cryst Sect D Struct Biol

Self Cite

View full text Add to dashboard Cite

The purification and three-dimensional crystallization of membrane proteins are commonly affected by a cumulation of pathologies that are less prevalent in their soluble counterparts. This may include severe anisotropy, poor spot shape, poor to moderate-resolution diffraction, crystal twinning, translational pseudo-symmetry and poor uptake of heavy atoms for derivatization. Such challenges must be circumvented by adaptations in the approach to crystallization and/or phasing. Here, an example of a protein that exhibited all of the above-mentioned complications is presented. Bestrophin-1 is a eukaryotic calcium-activated chloride channel, the structure of which was recently determined in complex with monoclonal antibody fragments using SAD phasing with tantalum bromide clusters (Ta6Br12·Br2). Some of the obstacles to obtaining improved diffraction and phasing for this particular channel are discussed, as well as the approach and adaptations that were key to determining the structure.

show abstract

Chloride Channels

Akabas¹

2020

Encyclopedia of Life Sciences

View full text Add to dashboard Cite

Chloride channels are integral membrane proteins that regulate the movement of chloride ions across cellular membranes. They perform a vital role in physiological processes such as epithelial transport, the regulation of nerve and muscle cell membrane excitability, cell volume regulation and in determining the pH within cytoplasmic membrane‐bound organelles. Several distinct protein families form chloride channels whose opening is regulated by different stimuli including neurotransmitter binding, changes in the concentrations of intracellular second messengers including cAMP and Ca 2+ , and phosphorylation. High‐resolution protein structures of many types of chloride channels have been solved by x‐ray crystallography and cryo‐electron microscopy. Mutations in genes encoding chloride channels impair their function and cause a variety of genetic diseases. Several clinically useful medicines act by modulating the activity of specific types of chloride channels. Key Concepts Chloride channels are ion channels that facilitate the movement of chloride ions across cell membranes. Chloride channels play fundamental roles in diverse physiological processes. Several distinct gene families encode proteins that function as chloride channels and have unique atomic structures. Genetic diseases result due to mutations in chloride channels that impair their function. The mechanism of action of a number of drugs used clinically involves chloride channels as specific targets.

show abstract

Structure and insights into the function of a Ca2+-activated Cl− channel

Cited by 185 publications

References 61 publications

The lysosomal potassium channel TMEM175 adopts a novel tetrameric architecture

The lysosomal potassium channel TMEM175 adopts a novel tetrameric architecture

Phasing and structure of bestrophin-1: a case study in the use of heavy-atom cluster compounds with multi-subunit transmembrane proteins

Chloride Channels

Contact Info

Product

Resources

About