Cryo-electron microscopy revealed TACAN is extensively and specifically associated with membrane lipids

TACAN is not a mechanosensitive ion channel but significantly linked to the mechanical hyperalgesia. In this study, we show that the human TACAN is a homodimer with each monomer consisting of a body, a spring and a blade domains. The body domain contains six transmembrane helices that forms an independent channel. The spring domain adapts a loop-helix-loop configuration with the helix running within and parallel to the membrane. The blade domain is composed of two cytoplasmic helices. In addition, we found that all the helices of the body and the spring domains are specifically associated with membrane lipids. Particularly, a lipid core, residing within a cavity formed by the two body and spring domains, contacts with the helices from the body and spring domains and extends to reach two symmetrically arranged lipid clusters. These results extremely imply that the membrane lipids coordinate with the membrane-embedded protein to sense and transduce the mechanic signal.

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Structure of the human epithelial sodium channel by cryo-electron microscopy

eLife ◽

10.7554/elife.39340 ◽

2018 ◽

Vol 7 ◽

Cited By ~ 47

Author(s):

Sigrid Noreng ◽

Arpita Bharadwaj ◽

Richard Posert ◽

Craig Yoshioka ◽

Isabelle Baconguis

Keyword(s):

Electron Microscopy ◽

Ion Channel ◽

Sodium Channel ◽

Conformational Changes ◽

Single Particle ◽

Transmembrane Domain ◽

Epithelial Sodium Channel ◽

Cryo Electron Microscopy ◽

Inhibitory Domain ◽

Epithelial Sodium Channel Enac

The epithelial sodium channel (ENaC), a member of the ENaC/DEG superfamily, regulates Na+ and water homeostasis. ENaCs assemble as heterotrimeric channels that harbor protease-sensitive domains critical for gating the channel. Here, we present the structure of human ENaC in the uncleaved state determined by single-particle cryo-electron microscopy. The ion channel is composed of a large extracellular domain and a narrow transmembrane domain. The structure reveals that ENaC assembles with a 1:1:1 stoichiometry of α:β:γ subunits arranged in a counter-clockwise manner. The shape of each subunit is reminiscent of a hand with key gating domains of a ‘finger’ and a ‘thumb.’ Wedged between these domains is the elusive protease-sensitive inhibitory domain poised to regulate conformational changes of the ‘finger’ and ‘thumb’; thus, the structure provides the first view of the architecture of inhibition of ENaC.

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Cryo-electron microscopy structure of a nucleosome-bound SWI/SNF chromatin remodeling complex

10.1101/805184 ◽

2019 ◽

Cited By ~ 2

Author(s):

Yan Han ◽

Alexis A Reyes ◽

Sara Malik ◽

Yuan He

Keyword(s):

Electron Microscopy ◽

Chromatin Remodeling ◽

Atp Hydrolysis ◽

Protein Complexes ◽

Gene Activation ◽

The Body ◽

Cellular Processes ◽

Cryo Electron Microscopy ◽

Chromatin Remodeling Complex ◽

High Resolution Structure

AbstractThe multi-subunit chromatin remodeling complex SWI/SNF1–3 is highly conserved from yeast to humans and plays critical roles in various cellular processes including transcription and DNA damage repair4, 5. It uses the energy from ATP hydrolysis to remodel chromatin structure by sliding and evicting the histone octamer6–10, creating DNA regions that become accessible to other essential protein complexes. However, our mechanistic understanding of the chromatin remodeling activity is largely hindered by the lack of a high-resolution structure of any complex from this family. Here we report the first structure of SWI/SNF from the yeast S. cerevisiae bound to a nucleosome at near atomic resolution determined by cryo-electron microscopy (cryo-EM). In the structure, the Arp module is sandwiched between the ATPase and the Body module of the complex, with the Snf2 HSA domain connecting all modules. The HSA domain also extends into the Body and anchors at the opposite side of the complex. The Body contains an assembly scaffold composed of conserved subunits Snf12 (SMARCD/BAF60), Snf5 (SMARCB1/BAF47/ INI1) and an asymmetric dimer of Swi3 (SMARCC/BAF155/170). Another conserved subunit Swi1 (ARID1/BAF250) folds into an Armadillo (ARM) repeat domain that resides in the core of the SWI/SNF Body, acting as a molecular hub. In addition to the interaction between Snf2 and the nucleosome, we also observed interactions between the conserved Snf5 subunit and the histones at the acidic patch, which could serve as an anchor point during active DNA translocation. Our structure allows us to map and rationalize a subset of cancer-related mutations in the human SWI/SNF complex and propose a model of how SWI/SNF recognizes and remodels the +1 nucleosome to generate nucleosome-depleted regions during gene activation11–13.

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Heterogeneous Populations of Ligand-Gated Ion Channel, GLIC, Revealed by Cryo-Electron Microscopy and Simulations

Biophysical Journal ◽

10.1016/j.bpj.2020.11.1321 ◽

2021 ◽

Vol 120 (3) ◽

pp. 192a

Author(s):

Urska Rovsnik ◽

Yuxuan Zhuang ◽

Bjorn Forsberg ◽

Marta Carroni ◽

Linnea Axelsson ◽

...

Keyword(s):

Electron Microscopy ◽

Ion Channel ◽

Heterogeneous Populations ◽

Cryo Electron Microscopy

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Structure-based identification and characterisation of novel inhibitors of KNa1.1 potassium channels, a stratified target for KCNT1-related epilepsy

10.1101/779975 ◽

2019 ◽

Author(s):

Bethan A. Cole ◽

Rachel M. Johnson ◽

Hattapark Dejakaisaya ◽

Nadia Pilati ◽

Colin W.G. Fishwick ◽

...

Keyword(s):

Electron Microscopy ◽

Drug Discovery ◽

Ion Channel ◽

Potassium Channel ◽

Mutational Analysis ◽

Drug Resistant ◽

Channel Activity ◽

Epileptic Encephalopathies ◽

Cytotoxicity Assays ◽

Cryo Electron Microscopy

AbstractSeveral types of drug-resistant epileptic encephalopathies of infancy have been associated with mutations in the KCNT1 gene, which encodes the sodium-activated potassium channel subunit KNa1.1. These mutations are commonly gain-of-function, increasing channel activity, therefore inhibition by drugs is proposed as a stratified approach to treat disorders. To date, quinidine therapy has been trialled with several patients, but mostly with unsuccessful outcomes, which has been linked to its low potency and lack of specificity. Here we describe the use of a cryo-electron microscopy-derived KNa1.1 structure and mutational analysis to identify the quinidine biding site and identified novel inhibitors that target this site using computational methods. We describe six compounds that inhibit KNa1.1 channels with low- and sub-micromolar potencies, likely through binding in the intracellular pore vestibule. In preliminary hERG inhibition and cytotoxicity assays, two compounds showed little effect. These compounds may provide starting points for the development of novel pharmacophores for KNa1.1 inhibition, with the view to treating KCNT1-associated epilepsy and, with their potencies higher than quinidine, could become key tool compounds to further study this channel. Furthermore, this study illustrates the potential for utilising cryo-electron microscopy in ion channel drug discovery.

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Faculty Opinions recommendation of Structure of a CLC chloride ion channel by cryo-electron microscopy.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.727132386.793528656 ◽

2017 ◽

Author(s):

Richard J Naftalin

Keyword(s):

Electron Microscopy ◽

Ion Channel ◽

Chloride Ion ◽

Cryo Electron Microscopy

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Cryo-electron microscopy structure of the TRPV2 ion channel

Nature Structural & Molecular Biology ◽

10.1038/nsmb.3159 ◽

2016 ◽

Vol 23 (2) ◽

pp. 180-186 ◽

Cited By ~ 177

Author(s):

Lejla Zubcevic ◽

Mark A Herzik ◽

Ben C Chung ◽

Zhirui Liu ◽

Gabriel C Lander ◽

...

Keyword(s):

Electron Microscopy ◽

Ion Channel ◽

Cryo Electron Microscopy

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Comparison of the transmembrane helices of bovine rhodopsin in the crystal structure and the C α template based on cryo-electron microscopy maps and sequence analysis of the G protein-coupled receptors

Molecular Simulation ◽

10.1080/0892702021000002520 ◽

2002 ◽

Vol 28 (8-9) ◽

pp. 845-851 ◽

Cited By ~ 2

Author(s):

Siavoush Dastmalchi ◽

Felix J. Kobus ◽

Tiina P. Iismaa ◽

Michael B. Morris ◽

W. Bret Church

Keyword(s):

Crystal Structure ◽

Electron Microscopy ◽

Sequence Analysis ◽

G Protein ◽

G Protein Coupled Receptors ◽

Transmembrane Helices ◽

Cryo Electron Microscopy ◽

Bovine Rhodopsin ◽

G Protein Coupled

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Stepwise gating of the Sec61 protein-conducting channel by Sec63 and Sec62

10.1101/2020.12.14.422728 ◽

2020 ◽

Author(s):

Samuel Itskanov ◽

Eunyong Park

Keyword(s):

Electron Microscopy ◽

Transmembrane Helices ◽

Conducting Channel ◽

Essential Step ◽

Gating Mechanisms ◽

Cryo Electron Microscopy ◽

Gate Opening ◽

Lateral Gate ◽

Sec61 Channel ◽

Hierarchical Manner

SummaryThe universally conserved Sec61/SecY channel mediates transport of many newly synthesized polypeptides across membranes, an essential step in protein secretion and membrane protein integration1-5. The channel has two gating mechanisms—a lipid-facing lateral gate, through which hydrophobic signal sequences or transmembrane helices (TMs) are released into the membrane, and a vertical gate, called the plug, which regulates the water-filled pore required for translocation of hydrophilic polypeptide segments6. Currently, how these gates are controlled and how they regulate the translocation process remain poorly understood. Here, by analyzing cryo-electron microscopy (cryo-EM) structures of several variants of the eukaryotic post-translational translocation complex Sec61-Sec62-Sec63, we reveal discrete gating steps of Sec61 and the mechanism by which Sec62 and Sec63 induce these gating events. We show that Sec62 forms a V-shaped structure in front of the lateral gate to fully open both gates of Sec61. Without Sec62, the lateral gate opening narrows, and the vertical pore becomes closed by the plug, rendering the channel inactive. We further show that the lateral gate is opened first by interactions between Sec61 and Sec63 in both cytosolic and luminal domains, a simultaneous disruption of which fully closes the channel. Our study defines the function of Sec62 and illuminates how Sec63 and Sec62 work together in a hierarchical manner to activate the Sec61 channel for post-translational translocation.

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