scholarly journals Cryo-EM structures of calcium homeostasis modulator channels in diverse oligomeric assemblies

2020 ◽  
Vol 6 (29) ◽  
pp. eaba8105 ◽  
Author(s):  
Kanae Demura ◽  
Tsukasa Kusakizako ◽  
Wataru Shihoya ◽  
Masahiro Hiraizumi ◽  
Kengo Nomura ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Caenorhabditis Elegans ◽  
Adenosine Triphosphate ◽  
Calcium Homeostasis ◽  
Transmembrane Domain ◽  
Atp Release ◽  
Channel Pore ◽  
Cryo Electron Microscopy ◽  
Chimeric Construct ◽  
Intersubunit Interactions

Calcium homeostasis modulator (CALHM) family proteins are Ca2+-regulated adenosine triphosphate (ATP)–release channels involved in neural functions including neurotransmission in gustation. Here, we present the cryo–electron microscopy (EM) structures of killifish CALHM1, human CALHM2, and Caenorhabditis elegans CLHM-1 at resolutions of 2.66, 3.4, and 3.6 Å, respectively. The CALHM1 octamer structure reveals that the N-terminal helix forms the constriction site at the channel pore in the open state and modulates the ATP conductance. The CALHM2 undecamer and CLHM-1 nonamer structures show the different oligomeric stoichiometries among CALHM homologs. We further report the cryo-EM structures of the chimeric construct, revealing that the intersubunit interactions at the transmembrane domain (TMD) and the TMD–intracellular domain linker define the oligomeric stoichiometry. These findings advance our understanding of the ATP conduction and oligomerization mechanisms of CALHM channels.

scholarly journals Cryo-EM structures of calcium homeostasis modulator channels in diverse oligomeric assemblies

2020 ◽  
Author(s):  
Kanae Demura ◽  
Tsukasa Kusakizako ◽  
Wataru Shihoya ◽  
Masahiro Hiraizumi ◽  
Kengo Nomura ◽  
...  
Keyword(s):  
Calcium Homeostasis ◽  
Transmembrane Domain ◽  
Atp Release ◽  
Subunit Interactions ◽  
Channel Pore ◽  
C Elegans ◽  
Oligomeric Assembly ◽  
Chimeric Construct

AbstractCalcium homeostasis modulator (CALHM) family proteins are Ca2+-regulated ATP-release channels involved in neural functions including neurotransmission in gustation. Here we present the cryo-EM structures of killifish CALHM1, human CALHM2, and C. elegans CLHM-1 at resolutions of 2.66, 3.51, and 3.60 Å, respectively. The CALHM1 octamer structure reveals that the N-terminal helix forms the constriction site at the channel pore in the open state, and modulates the ATP conductance. The CALHM2 undecamer and CLHM-1 nonomer structures show the different oligomeric stoichiometries among CALHM homologs. We further report the cryo-EM structures of the chimeric construct, revealing that the inter-subunit interactions at the transmembrane domain define the oligomeric stoichiometry. These findings advance our understanding of the ATP conduction and oligomerization mechanisms of CALHM channels.One Sentence SummaryCryo-EM structures reveal the ATP conduction and oligomeric assembly mechanisms of CALHM channels.

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

eLife ◽  
2018 ◽  
Vol 7 ◽  
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.

scholarly journals Cryo-EM structure of the calcium homeostasis modulator 1 channel

2020 ◽  
Vol 6 (29) ◽  
pp. eaba8161
Author(s):  
Yue Ren ◽  
Tianlei Wen ◽  
Zhiqin Xi ◽  
Shunjin Li ◽  
Jing Lu ◽  
...  
Keyword(s):  
Calcium Homeostasis ◽  
Transmembrane Helix ◽  
Atp Release ◽  
Ion Permeation ◽  
Extracellular Loop ◽  
Release Channel ◽  
Dimer Interface ◽  
Cryo Electron Microscopy ◽  
Gating Mechanism ◽  
Oligomeric Assembly

Calcium homeostasis modulator 1 (CALHM1) is a voltage-gated ATP release channel that plays an important role in neural gustatory signaling and the pathogenesis of Alzheimer’s disease. Here, we present a cryo–electron microscopy structure of full-length Ca2+-free CALHM1 from Danio rerio at an overall resolution of 3.1 Å. Our structure reveals an octameric architecture with a wide pore diameter of ~20 Å, presumably representing the active conformation. The overall structure is substantially different from that of the isoform CALHM2, which forms both undecameric hemichannels and gap junctions. The N-terminal small helix folds back to the pore and forms an antiparallel interaction with transmembrane helix 1. Structural analysis revealed that the extracellular loop 1 region within the dimer interface may contribute to oligomeric assembly. A positive potential belt inside the pore was identified that may modulate ion permeation. Our structure offers insights into the assembly and gating mechanism of the CALHM1 channel.

scholarly journals Cryo-EM structures of SERCA2b reveal the mechanism of regulation by the luminal extension tail

2020 ◽  
Vol 6 (33) ◽  
pp. eabb0147 ◽  
Author(s):  
Yuxia Zhang ◽  
Michio Inoue ◽  
Akihisa Tsutsumi ◽  
Satoshi Watanabe ◽  
Tomohiro Nishizawa ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Calcium Homeostasis ◽  
Critical Role ◽  
Kinetic Properties ◽  
Cryo Electron Microscopy ◽  
Domain Arrangement ◽  
Tm Domain ◽  
Conformational Regulation ◽  
Multiple Conformations

Sarco/endoplasmic reticulum Ca2+ ATPase (SERCA) pumps Ca2+ from the cytosol into the ER and maintains the cellular calcium homeostasis. Herein, we present cryo–electron microscopy (cryo-EM) structures of human SERCA2b in E1∙2Ca2+–adenylyl methylenediphosphonate (AMPPCP) and E2-BeF3− states at 2.9- and 2.8-Å resolutions, respectively. The structures revealed that the luminal extension tail (LE) characteristic of SERCA2b runs parallel to the lipid-water boundary near the luminal ends of transmembrane (TM) helices TM10 and TM7 and approaches the luminal loop flanked by TM7 and TM8. While the LE served to stabilize the cytosolic and TM domain arrangement of SERCA2b, deletion of the LE rendered the overall conformation resemble that of SERCA1a and SERCA2a and allowed multiple conformations. Thus, the LE appears to play a critical role in conformational regulation in SERCA2b, which likely explains the different kinetic properties of SERCA2b from those of other isoforms lacking the LE.

Structure and dynamics of the yeast SWR1-nucleosome complex

Science ◽  
2018 ◽  
Vol 362 (6411) ◽  
pp. eaat7716 ◽  
Author(s):  
Oliver Willhoft ◽  
Mohamed Ghoneim ◽  
Chia-Liang Lin ◽  
Eugene Y. D. Chua ◽  
Martin Wilkinson ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Adenosine Triphosphate ◽  
Single Molecule ◽  
Conformational Changes ◽  
Base Pair ◽  
Histone H2a ◽  
Substrate Interactions ◽  
Structure And Dynamics ◽  
Cryo Electron Microscopy ◽  
Histone Core

The yeast SWR1 complex exchanges histone H2A in nucleosomes with Htz1 (H2A.Z in humans). The cryo–electron microscopy structure of the SWR1 complex bound to a nucleosome at 3.6-angstrom resolution reveals details of the intricate interactions between components of the SWR1 complex and its nucleosome substrate. Interactions between the Swr1 motor domains and the DNA wrap at superhelical location 2 distort the DNA, causing a bulge with concomitant translocation of the DNA by one base pair, coupled to conformational changes of the histone core. Furthermore, partial unwrapping of the DNA from the histone core takes place upon binding of nucleosomes to SWR1 complex. The unwrapping, as monitored by single-molecule data, is stabilized and has its dynamics altered by adenosine triphosphate binding but does not require hydrolysis.

scholarly journals ATP and Substrate Binding Regulates Conformational Changes of Human Peroxisomal ABC Transporter ALDP

2021 ◽  
Author(s):  
Lin Tang ◽  
Chao Xiong ◽  
Lina Jia ◽  
Ming-He Shen ◽  
Wei-Xi Xiong ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Fatty Acids ◽  
Conformational Changes ◽  
Transmembrane Domain ◽  
Long Chain Fatty Acids ◽  
Variable Size ◽  
Binding Domains ◽  
Cryo Electron Microscopy ◽  
Peroxisome Membrane ◽  
Long Chain

Abstract The malfunction of ABCD1 causes X-linked adrenoleukodystrophy (X-ALD), a rare neurodegenerative disease that affect all tissues in human. Residing in the peroxisome membrane, ABCD1 plays a role in the translocation of very long chain fatty acids (VLCFA) for their damage by β-oxidation. Here, we present five Cryo-Electron microscopy structures of ABCD1 in four conformational states. Combined with functional analysis, we found that substrate and ATP trigger the closing of two nucleotide binding domains (NBDs) over a distance of 40 Å and the rearrangement of the transmembrane domains. Each of the three inward-facing structure of ABCD1 has a vestibule opens to cytosol with variable size. Furthermore, the structure of ABCD1 in the outward-facing state supports that ATP molecules pull the two NBDs together and open the transmembrane domain to the peroxisomal lumen for substrate release. The five structures provide a snapshot of substrate transporting cycle and mechanistic implications for disease-causing mutations.

scholarly journals Cryo-EM structures and transport mechanism of human P5B type ATPase ATP13A2

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Xudong Chen ◽  
Mingze Zhou ◽  
Sensen Zhang ◽  
Jian Yin ◽  
Ping Zhang ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Neurodegenerative Disorders ◽  
Mammalian Cells ◽  
Transport Mechanism ◽  
Transmembrane Domain ◽  
Normal Function ◽  
Cryo Electron Microscopy ◽  
Polyamine Transport ◽  
Adenosine Triphosphatases ◽  
Transport Cycle

AbstractPolyamines are important polycations that play critical roles in mammalian cells. ATP13A2 belongs to the orphan P5B adenosine triphosphatases (ATPase) family and has been established as a lysosomal polyamine exporter to maintain the normal function of lysosomes and mitochondria. Previous studies have reported that several human neurodegenerative disorders are related to mutations in the ATP13A2 gene. However, the transport mechanism of ATP13A2 in the lysosome remains unclear. Here, we report the cryo-electron microscopy (cryo-EM) structures of three distinct intermediates of the human ATP13A2, revealing key insights into the spermine (SPM) transport cycle in the lysosome. The transmembrane domain serves as a substrate binding site and the C-terminal domain is essential for protein stability and may play a regulatory role. These findings advance our understanding of the polyamine transport mechanism, the lipid-associated regulation, and the disease-associated mutants of ATP13A2.

scholarly journals Mechanism of CFTR correction by type I folding correctors

2021 ◽  
Author(s):  
Jue Chen ◽  
Karol Fiedorczuk
Keyword(s):  
Electron Microscopy ◽  
Cystic Fibrosis ◽  
Mechanism Of Action ◽  
Protein Misfolding ◽  
Transmembrane Domain ◽  
Early Stage ◽  
Type I ◽  
Hydrophobic Pocket ◽  
Cryo Electron Microscopy ◽  
Δf508 Cftr

Small molecule chaperones have been exploited as therapeutics for the hundreds of diseases caused by protein misfolding. The most successful examples are the CFTR correctors, which transformed cystic fibrosis therapy. These molecules revert folding defects of the ΔF508 mutant and are widely used to treat patients. However, their mechanism of action is unknown. Here we present cryo-electron microscopy structures of CFTR in complex with two FDA-approved correctors: lumacaftor and tezacaftor. Both drugs insert into a hydrophobic pocket in the first transmembrane domain (TMD1), linking together four helices that are thermodynamically unstable. Mutating residues at the binding site rendered ΔF508-CFTR insensitive to lumacaftor and tezacaftor, underscoring the functional significance of the structural discovery. These results support a mechanism in which the correctors stabilize TMD1 at an early stage of biogenesis, prevent its pre-mature degradation, and thereby allosterically rescue a large number of disease-causing mutations.

scholarly journals Cryo‐electron microscopy structure of CLHM1 ion channel from Caenorhabditis elegans

2020 ◽  
Vol 29 (8) ◽  
pp. 1803-1815
Author(s):  
Weixin Yang ◽  
Youwang Wang ◽  
Jianli Guo ◽  
Lingli He ◽  
Ye Zhou ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Caenorhabditis Elegans ◽  
Ion Channel ◽  
Cryo Electron Microscopy

scholarly journals ATP and Substrate Binding Regulates Conformational Changes of Human Peroxisomal ABC Transporter ALDP

2021 ◽  
Author(s):  
chao xiong ◽  
Li-Na Jia ◽  
Ming-He Shen ◽  
Wei-Xi Xiong ◽  
Liu-Lin Xiong ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Fatty Acids ◽  
Conformational Changes ◽  
Transmembrane Domain ◽  
Long Chain Fatty Acids ◽  
Variable Size ◽  
Binding Domains ◽  
Cryo Electron Microscopy ◽  
Peroxisome Membrane ◽  
Long Chain

The malfunction of ABCD1 causes X-linked adrenoleukodystrophy (X-ALD), a rare neurodegenerative disease that affect all tissues in human. Residing in the peroxisome membrane, ABCD1 plays a role in the translocation of very long chain fatty acids (VLCFA) for their damage by β-oxidation. Here, we present five Cryo-Electron microscopy structures of ABCD1 in four conformational states. Combined with functional analysis, we found that substrate and ATP trigger the closing of two nucleotide binding domains (NBDs) over a distance of 40 &Aring and the rearrangement of the transmembrane domains. Each of the three inward-facing structure of ABCD1 has a vestibule opens to cytosol with variable size. Furthermore, the structure of ABCD1 in the outward-facing state supports that ATP molecules pull the two NBDs together and open the transmembrane domain to the peroxisomal lumen for substrate release. The five structures provide a snapshot of substrate transporting cycle and mechanistic implications for disease-causing mutations.

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