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- EMDB-27436: Human TMEM175 in complex with 4-aminopyridine -

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Basic information

Entry
Database: EMDB / ID: EMD-27436
TitleHuman TMEM175 in complex with 4-aminopyridine
Map datahsTMEM175 in complex with 4-aminopyridine
Sample
  • Complex: Human TMEM175
    • Protein or peptide: Endosomal/lysosomal potassium channel TMEM175
  • Ligand: POTASSIUM ION
  • Ligand: 4-AMINOPYRIDINE
  • Ligand: water
Keywordspotassium channel / ion channel / TRANSPORT PROTEIN
Function / homology
Function and homology information


lysosomal lumen pH elevation / phagosome-lysosome fusion / regulation of lysosomal lumen pH / proton channel activity / potassium ion leak channel activity / arachidonic acid binding / potassium channel activity / potassium ion transmembrane transport / proton transmembrane transport / neuron cellular homeostasis ...lysosomal lumen pH elevation / phagosome-lysosome fusion / regulation of lysosomal lumen pH / proton channel activity / potassium ion leak channel activity / arachidonic acid binding / potassium channel activity / potassium ion transmembrane transport / proton transmembrane transport / neuron cellular homeostasis / lysosome / endosome membrane / endosome / lysosomal membrane
Similarity search - Function
Endosomal/lysomomal potassium channel TMEM175 / Endosomal/lysosomal potassium channel TMEM175
Similarity search - Domain/homology
Endosomal/lysosomal proton channel TMEM175
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.73 Å
AuthorsOh S / Hite RK
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM141553 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA008748 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2022
Title: Mechanism of 4-aminopyridine inhibition of the lysosomal channel TMEM175.
Authors: SeCheol Oh / Robyn Stix / Wenchang Zhou / José D Faraldo-Gómez / Richard K Hite /
Abstract: Transmembrane protein 175 (TMEM175) is an evolutionarily distinct lysosomal cation channel whose mutation is associated with the development of Parkinson's disease. Here, we present a cryoelectron ...Transmembrane protein 175 (TMEM175) is an evolutionarily distinct lysosomal cation channel whose mutation is associated with the development of Parkinson's disease. Here, we present a cryoelectron microscopy structure and molecular simulations of TMEM175 bound to 4-aminopyridine (4-AP), the only known small-molecule inhibitor of TMEM175 and a broad K channel inhibitor, as well as a drug approved by the Food and Drug Administration against multiple sclerosis. The structure shows that 4-AP, whose mode of action had not been previously visualized, binds near the center of the ion conduction pathway, in the open state of the channel. Molecular dynamics simulations reveal that this binding site is near the middle of the transmembrane potential gradient, providing a rationale for the voltage-dependent dissociation of 4-AP from TMEM175. Interestingly, bound 4-AP rapidly switches between three predominant binding poses, stabilized by alternate interaction patterns dictated by the twofold symmetry of the channel. Despite this highly dynamic binding mode, bound 4-AP prevents not only ion permeation but also water flow. Together, these studies provide a framework for the rational design of novel small-molecule inhibitors of TMEM175 that might reveal the role of this channel in human lysosomal physiology both in health and disease.
History
DepositionJun 27, 2022-
Header (metadata) releaseNov 23, 2022-
Map releaseNov 23, 2022-
UpdateJun 12, 2024-
Current statusJun 12, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27436.png

Downloads & links

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Map

FileDownload / File: emd_27436.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationhsTMEM175 in complex with 4-aminopyridine
Voxel sizeX=Y=Z: 0.85 Å
Density
Contour LevelBy AUTHOR: 0.4
Minimum - Maximum-0.7536181 - 1.6678687
Average (Standard dev.)-0.00014015174 (±0.030461138)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 326.40002 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Density modified and resampled 1.5x at

Fileemd_27436_additional_1.map
AnnotationDensity modified and resampled 1.5x at
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map 1

Fileemd_27436_half_map_1.map
AnnotationHalf Map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map 2

Fileemd_27436_half_map_2.map
AnnotationHalf Map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Human TMEM175

EntireName: Human TMEM175
Components
  • Complex: Human TMEM175
    • Protein or peptide: Endosomal/lysosomal potassium channel TMEM175
  • Ligand: POTASSIUM ION
  • Ligand: 4-AMINOPYRIDINE
  • Ligand: water

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Supramolecule #1: Human TMEM175

SupramoleculeName: Human TMEM175 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Endosomal/lysosomal potassium channel TMEM175

MacromoleculeName: Endosomal/lysosomal potassium channel TMEM175 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 55.667219 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MSQPRTPEQA LDTPGDCPPG RRDEDAGEGI QCSQRMLSFS DALLSIIATV MILPVTHTEI SPEQQFDRSV QRLLATRIAV YLMTFLIVT VAWAAHTRLF QVVGKTDDTL ALLNLACMMT ITFLPYTFSL MVTFPDVPLG IFLFCVCVIA IGVVQALIVG Y AFHFPHLL ...String:
MSQPRTPEQA LDTPGDCPPG RRDEDAGEGI QCSQRMLSFS DALLSIIATV MILPVTHTEI SPEQQFDRSV QRLLATRIAV YLMTFLIVT VAWAAHTRLF QVVGKTDDTL ALLNLACMMT ITFLPYTFSL MVTFPDVPLG IFLFCVCVIA IGVVQALIVG Y AFHFPHLL SPQIQRSAHR ALYRRHVLGI VLQGPALCFA AAIFSLFFVP LSYLLMVTVI LLPYVSKVTG WCRDRLLGHR EP SAHPVEV FSFDLHEPLS KERVEAFSDG VYAIVATLLI LDICEDNVPD PKDVKERFSG SLVAALSATG PRFLAYFGSF ATV GLLWFA HHSLFLHVRK ATRAMGLLNT LSLAFVGGLP LAYQQTSAFA RQPRDELERV RVSCTIIFLA SIFQLAMWTT ALLH QAETL QPSVWFGGRE HVLMFAKLAL YPCASLLAFA STCLLSRFSV GIFHLMQIAV PCAFLLLRLL VGLALATLRV LRGLA RPEH PPPAPTGQDD PQSQLLPAPC

UniProtKB: Endosomal/lysosomal proton channel TMEM175

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Macromolecule #2: POTASSIUM ION

MacromoleculeName: POTASSIUM ION / type: ligand / ID: 2 / Number of copies: 4 / Formula: K
Molecular weightTheoretical: 39.098 Da

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Macromolecule #3: 4-AMINOPYRIDINE

MacromoleculeName: 4-AMINOPYRIDINE / type: ligand / ID: 3 / Number of copies: 1 / Formula: 4AP
Molecular weightTheoretical: 95.122 Da
Chemical component information

ChemComp-4AP:
4-AMINOPYRIDINE

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 91 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration4 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMKClPotassium chloride
50.0 mMTris-HCl
0.1 %Lauryl maltose neopentyl glycol
2.0 mMDithiothreitol
1.0 %Dimethyl sulfoxide
1.0 mM4-aminopyridine
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 10 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 297 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 61.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 4153614
Startup modelType of model: INSILICO MODEL
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.73 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 261536
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationSoftware - Name: cryoSPARC
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6wc9


Chain - Source name: PDB / Chain - Initial model type: experimental model
DetailsReal Space Refinement
RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-8dhm:
Human TMEM175 in complex with 4-aminopyridine

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