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- EMDB-22389: Structure of human CLC-7/OSTM1 complex -

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Basic information

Entry
Database: EMDB / ID: EMD-22389
TitleStructure of human CLC-7/OSTM1 complex
Map dataDensity map of human CLC-7/OSTM1
Sample
  • Complex: Human CLC-7/OSTM1 complex
    • Protein or peptide: Osteopetrosis-associated transmembrane protein 1
    • Protein or peptide: H(+)/Cl(-) exchange transporter 7
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: CHLORIDE IONChloride
  • Ligand: (2R)-3-{[(R)-hydroxy{[(1S,2R,3S,4S,5R,6R)-2,3,4,6-tetrahydroxy-5-(phosphonooxy)cyclohexyl]oxy}phosphoryl]oxy}propane-1,2-diyl dinonanoate
  • Ligand: water
Function / homology
Function and homology information


transepithelial chloride transport / voltage-gated chloride channel activity / antiporter activity / chloride transmembrane transporter activity / chloride channel activity / chloride channel complex / osteoclast differentiation / Stimuli-sensing channels / lysosomal membrane / intracellular membrane-bounded organelle ...transepithelial chloride transport / voltage-gated chloride channel activity / antiporter activity / chloride transmembrane transporter activity / chloride channel activity / chloride channel complex / osteoclast differentiation / Stimuli-sensing channels / lysosomal membrane / intracellular membrane-bounded organelle / ATP binding / membrane / cytosol
Similarity search - Function
Osteopetrosis-associated transmembrane protein 1 precursor / Osteopetrosis-associated transmembrane protein 1 precursor / Chloride channel ClC-7 / Chloride channel, voltage gated / Chloride channel, core / Voltage gated chloride channel / CBS domain superfamily / Domain in cystathionine beta-synthase and other proteins. / CBS domain / CBS domain / CBS domain profile.
Similarity search - Domain/homology
H(+)/Cl(-) exchange transporter 7 / Osteopetrosis-associated transmembrane protein 1
Similarity search - Component
Biological speciesHomo (humans) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.82 Å
AuthorsSchrecker M / Hite R
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103310 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)P30 CA008748 United States
CitationJournal: Elife / Year: 2020
Title: Cryo-EM structure of the lysosomal chloride-proton exchanger CLC-7 in complex with OSTM1.
Authors: Marina Schrecker / Julia Korobenko / Richard K Hite /
Abstract: The chloride-proton exchanger CLC-7 plays critical roles in lysosomal homeostasis and bone regeneration and its mutation can lead to osteopetrosis, lysosomal storage disease and neurological ...The chloride-proton exchanger CLC-7 plays critical roles in lysosomal homeostasis and bone regeneration and its mutation can lead to osteopetrosis, lysosomal storage disease and neurological disorders. In lysosomes and the ruffled border of osteoclasts, CLC-7 requires a β-subunit, OSTM1, for stability and activity. Here, we present electron cryomicroscopy structures of CLC-7 in occluded states by itself and in complex with OSTM1, determined at resolutions up to 2.8 Å. In the complex, the luminal surface of CLC-7 is entirely covered by a dimer of the heavily glycosylated and disulfide-bonded OSTM1, which serves to protect CLC-7 from the degradative environment of the lysosomal lumen. OSTM1 binding does not induce large-scale rearrangements of CLC-7, but does have minor effects on the conformation of the ion-conduction pathway, potentially contributing to its regulatory role. These studies provide insights into the role of OSTM1 and serve as a foundation for understanding the mechanisms of CLC-7 regulation.
History
DepositionJul 31, 2020-
Header (metadata) releaseSep 2, 2020-
Map releaseSep 2, 2020-
UpdateSep 2, 2020-
Current statusSep 2, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 2.5
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 2.5
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7jm7
  • Surface level: 2.5
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22389.png

Downloads & links

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Map

FileDownload / File: emd_22389.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationDensity map of human CLC-7/OSTM1
Voxel sizeX=Y=Z: 0.70933 Å
Density
Contour LevelBy AUTHOR: 2.5 / Movie #1: 2.5
Minimum - Maximum-42.712944 - 60.458668
Average (Standard dev.)-0.0012214383 (±1.143893)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 272.38388 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.709333333333330.709333333333330.70933333333333
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z272.384272.384272.384
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ384384384
MAP C/R/S321
start NC/NR/NS000
NC/NR/NS384384384
D min/max/mean-42.71360.459-0.001

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Supplemental data

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Sample components

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Entire : Human CLC-7/OSTM1 complex

EntireName: Human CLC-7/OSTM1 complex
Components
  • Complex: Human CLC-7/OSTM1 complex
    • Protein or peptide: Osteopetrosis-associated transmembrane protein 1
    • Protein or peptide: H(+)/Cl(-) exchange transporter 7
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: CHLORIDE IONChloride
  • Ligand: (2R)-3-{[(R)-hydroxy{[(1S,2R,3S,4S,5R,6R)-2,3,4,6-tetrahydroxy-5-(phosphonooxy)cyclohexyl]oxy}phosphoryl]oxy}propane-1,2-diyl dinonanoate
  • Ligand: water

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Supramolecule #1: Human CLC-7/OSTM1 complex

SupramoleculeName: Human CLC-7/OSTM1 complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Homo (humans)
Recombinant expressionOrganism: Homo sapiens (human)

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Macromolecule #1: Osteopetrosis-associated transmembrane protein 1

MacromoleculeName: Osteopetrosis-associated transmembrane protein 1 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 37.29057 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MEPGPTAAQR RCSLPPWLPL GLLLWSGLAL GALPFGSSPH RVFHDLLSEQ QLLEVEDLSL SLLQGGGLGP LSLPPDLPDL DPECRELLL DFANSSAELT GCLVRSARPV RLCQTCYPLF QQVVSKMDNI SRAAGNTSES QSCARSLLMA DRMQIVVILS E FFNTTWQE ...String:
MEPGPTAAQR RCSLPPWLPL GLLLWSGLAL GALPFGSSPH RVFHDLLSEQ QLLEVEDLSL SLLQGGGLGP LSLPPDLPDL DPECRELLL DFANSSAELT GCLVRSARPV RLCQTCYPLF QQVVSKMDNI SRAAGNTSES QSCARSLLMA DRMQIVVILS E FFNTTWQE ANCANCLTNN SEELSNSTVY FLNLFNHTLT CFEHNLQGNA HSLLQTKNYS EVCKNCREAY KTLSSLYSEM QK MNELENK AEPGTHLCID VEDAMNITRK LWSRTFNCSV PCSDTVPVIA VSVFILFLPV VFYLSSFLHS EQKKRKLILP KRL KSSTSF ANIQENSN

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Macromolecule #2: H(+)/Cl(-) exchange transporter 7

MacromoleculeName: H(+)/Cl(-) exchange transporter 7 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 88.773875 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MANVSKKVSW SGRDRDDEEA APLLRRTARP GGGTPLLNGA GPGAARQSPR SALFRVGHMS SVELDDELLD PDMDPPHPFP KEIPHNEKL LSLKYESLDY DNSENQLFLE EERRINHTAF RTVEIKRWVI CALIGILTGL VACFIDIVVE NLAGLKYRVI K GNIDKFTE ...String:
MANVSKKVSW SGRDRDDEEA APLLRRTARP GGGTPLLNGA GPGAARQSPR SALFRVGHMS SVELDDELLD PDMDPPHPFP KEIPHNEKL LSLKYESLDY DNSENQLFLE EERRINHTAF RTVEIKRWVI CALIGILTGL VACFIDIVVE NLAGLKYRVI K GNIDKFTE KGGLSFSLLL WATLNAAFVL VGSVIVAFIE PVAAGSGIPQ IKCFLNGVKI PHVVRLKTLV IKVSGVILSV VG GLAVGKE GPMIHSGSVI AAGISQGRST SLKRDFKIFE YFRRDTEKRD FVSAGAAAGV SAAFGAPVGG VLFSLEEGAS FWN QFLTWR IFFASMISTF TLNFVLSIYH GNMWDLSSPG LINFGRFDSE KMAYTIHEIP VFIAMGVVGG VLGAVFNALN YWLT MFRIR YIHRPCLQVI EAVLVAAVTA TVAFVLIYSS RDCQPLQGGS MSYPLQLFCA DGEYNSMAAA FFNTPEKSVV SLFHD PPGS YNPLTLGLFT LVYFFLACWT YGLTVSAGVF IPSLLIGAAW GRLFGISLSY LTGAAIWADP GKYALMGAAA QLGGIV RMT LSLTVIMMEA TSNVTYGFPI MLVLMTAKIV GDVFIEGLYD MHIQLQSVPF LHWEAPVTSH SLTAREVMST PVTCLRR RE KVGVIVDVLS DTASNHNGFP VVEHADDTQP ARLQGLILRS QLIVLLKHKV FVERSNLGLV QRRLRLKDFR DAYPRFPP I QSIHVSQDER ECTMDLSEFM NPSPYTVPQE ASLPRVFKLF RALGLRHLVV VDNRNQVVGL VTRKDLARYR LGKRGLEEL SLAQT

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Macromolecule #5: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 5 / Number of copies: 8 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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Macromolecule #6: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 6 / Number of copies: 2 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM / Adenosine triphosphate

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Macromolecule #7: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 7 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #8: CHLORIDE ION

MacromoleculeName: CHLORIDE ION / type: ligand / ID: 8 / Number of copies: 6 / Formula: CL
Molecular weightTheoretical: 35.453 Da

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Macromolecule #9: (2R)-3-{[(R)-hydroxy{[(1S,2R,3S,4S,5R,6R)-2,3,4,6-tetrahydroxy-5-...

MacromoleculeName: (2R)-3-{[(R)-hydroxy{[(1S,2R,3S,4S,5R,6R)-2,3,4,6-tetrahydroxy-5-(phosphonooxy)cyclohexyl]oxy}phosphoryl]oxy}propane-1,2-diyl dinonanoate
type: ligand / ID: 9 / Number of copies: 2 / Formula: 0J1
Molecular weightTheoretical: 694.64 Da
Chemical component information

ChemComp-0J1:
(2R)-3-{[(R)-hydroxy{[(1S,2R,3S,4S,5R,6R)-2,3,4,6-tetrahydroxy-5-(phosphonooxy)cyclohexyl]oxy}phosphoryl]oxy}propane-1,2-diyl dinonanoate

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Macromolecule #10: water

MacromoleculeName: water / type: ligand / ID: 10 / Number of copies: 36 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER / 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

Concentration2 mg/mL
BufferpH: 8
Component:
ConcentrationFormula
150.0 mMKCL
50.0 mMTris
2.0 mMDTT
0.1 %LMNG
1.0 mMATPAdenosine triphosphate
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: OTHER / Cs: 2.7 mm / Nominal defocus max: -2.7 µm / Nominal defocus min: -1.2 µm / Nominal magnification: 22500
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Detector mode: SUPER-RESOLUTION / Average exposure time: 8.0 sec. / Average electron dose: 44.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: CTFFIND
Startup modelType of model: OTHER / Details: CryoSparc Ab-initio
Initial angle assignmentType: NOT APPLICABLE
Final 3D classificationNumber classes: 8 / Software - Name: cryoSPARC (ver. V2.15.0)
Final angle assignmentType: OTHER / Software - Name: cryoSPARC (ver. V2.15.0) / Details: CryoSparc Non-uniform refinement
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.82 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 327619

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: OTHER
Output model

PDB-7jm7:
Structure of human CLC-7/OSTM1 complex

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