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- EMDB-35399: Structure of human alpha-2/delta-1 with mirogabalin -

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Basic information

Entry
Database: EMDB / ID: EMD-35399
TitleStructure of human alpha-2/delta-1 with mirogabalin
Map data
Sample
  • Organelle or cellular component: Voltage-dependent calcium channel subunit alpha-2/delta-1
    • Protein or peptide: Voltage-dependent calcium channel subunit alpha-2/delta-1
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: 2-[(1R,5S,6S)-6-(aminomethyl)-3-ethyl-6-bicyclo[3.2.0]hept-3-enyl]acetic acid
Function / homology
Function and homology information


regulation of membrane repolarization during action potential / positive regulation of high voltage-gated calcium channel activity / calcium ion transmembrane transport via high voltage-gated calcium channel / membrane depolarization during bundle of His cell action potential / L-type voltage-gated calcium channel complex / cardiac muscle cell action potential involved in contraction / regulation of ventricular cardiac muscle cell membrane repolarization / calcium ion transport into cytosol / regulation of calcium ion transmembrane transport via high voltage-gated calcium channel / voltage-gated calcium channel complex ...regulation of membrane repolarization during action potential / positive regulation of high voltage-gated calcium channel activity / calcium ion transmembrane transport via high voltage-gated calcium channel / membrane depolarization during bundle of His cell action potential / L-type voltage-gated calcium channel complex / cardiac muscle cell action potential involved in contraction / regulation of ventricular cardiac muscle cell membrane repolarization / calcium ion transport into cytosol / regulation of calcium ion transmembrane transport via high voltage-gated calcium channel / voltage-gated calcium channel complex / calcium ion import across plasma membrane / neuronal dense core vesicle / regulation of heart rate by cardiac conduction / regulation of calcium ion transport / voltage-gated calcium channel activity / sarcoplasmic reticulum / cellular response to amyloid-beta / calcium ion transport / extracellular exosome / metal ion binding / plasma membrane
Similarity search - Function
VWA N-terminal / Voltage-dependent calcium channel, alpha-2/delta subunit, conserved region / : / VWA N-terminal / Neuronal voltage-dependent calcium channel alpha 2acd / von Willebrand factor type A domain / von Willebrand factor (vWF) type A domain / VWFA domain profile. / von Willebrand factor, type A / von Willebrand factor A-like domain superfamily
Similarity search - Domain/homology
Voltage-dependent calcium channel subunit alpha-2/delta-1
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.23 Å
AuthorsKozai D / Numoto N / Fujiyoshi Y
Funding support Japan, 2 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)20H00451 Japan
Japan Agency for Medical Research and Development (AMED) Japan
CitationJournal: J Mol Biol / Year: 2023
Title: Recognition Mechanism of a Novel Gabapentinoid Drug, Mirogabalin, for Recombinant Human αδ1, a Voltage-Gated Calcium Channel Subunit.
Authors: Daisuke Kozai / Nobutaka Numoto / Kouki Nishikawa / Akiko Kamegawa / Shohei Kawasaki / Yoko Hiroaki / Katsumasa Irie / Atsunori Oshima / Hiroyuki Hanzawa / Kousei Shimada / Yutaka Kitano / Yoshinori Fujiyoshi /
Abstract: Mirogabalin is a novel gabapentinoid drug with a hydrophobic bicyclo substituent on the γ-aminobutyric acid moiety that targets the voltage-gated calcium channel subunit αδ1. Here, to reveal the ...Mirogabalin is a novel gabapentinoid drug with a hydrophobic bicyclo substituent on the γ-aminobutyric acid moiety that targets the voltage-gated calcium channel subunit αδ1. Here, to reveal the mirogabalin recognition mechanisms of αδ1, we present structures of recombinant human αδ1 with and without mirogabalin analyzed by cryo-electron microscopy. These structures show the binding of mirogabalin to the previously reported gabapentinoid binding site, which is the extracellular dCache_1 domain containing a conserved amino acid binding motif. A slight conformational change occurs around the residues positioned close to the hydrophobic group of mirogabalin. Mutagenesis binding assays identified that residues in the hydrophobic interaction region, in addition to several amino acid binding motif residues around the amino and carboxyl groups of mirogabalin, are critical for mirogabalin binding. The A215L mutation introduced to decrease the hydrophobic pocket volume predictably suppressed mirogabalin binding and promoted the binding of another ligand, L-Leu, with a smaller hydrophobic substituent than mirogabalin. Alterations of residues in the hydrophobic interaction region of αδ1 to those of the αδ2, αδ3, and αδ4 isoforms, of which αδ3 and αδ4 are gabapentin-insensitive, suppressed the binding of mirogabalin. These results support the importance of hydrophobic interactions in αδ1 ligand recognition.
History
DepositionFeb 17, 2023-
Header (metadata) releaseApr 5, 2023-
Map releaseApr 5, 2023-
UpdateApr 12, 2023-
Current statusApr 12, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_35399.png

Downloads & links

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Map

FileDownload / File: emd_35399.map.gz / Format: CCP4 / Size: 27.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.77 Å/pix.
x 194 pix.
= 148.41 Å		0.77 Å/pix.
x 194 pix.
= 148.41 Å		0.77 Å/pix.
x 194 pix.
= 148.41 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.765 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0077
Minimum - Maximum-0.03978311 - 0.053114537
Average (Standard dev.)-2.0547224e-05 (±0.0022363132)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions194194194
Spacing194194194
CellA=B=C: 148.41 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_35399_msk_1.map
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Half map: #1

Fileemd_35399_half_map_1.map
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Half map: #2

Fileemd_35399_half_map_2.map
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Sample components

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Entire : Voltage-dependent calcium channel subunit alpha-2/delta-1

EntireName: Voltage-dependent calcium channel subunit alpha-2/delta-1
Components
  • Organelle or cellular component: Voltage-dependent calcium channel subunit alpha-2/delta-1
    • Protein or peptide: Voltage-dependent calcium channel subunit alpha-2/delta-1
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
  • Ligand: 2-[(1R,5S,6S)-6-(aminomethyl)-3-ethyl-6-bicyclo[3.2.0]hept-3-enyl]acetic acid

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Supramolecule #1: Voltage-dependent calcium channel subunit alpha-2/delta-1

SupramoleculeName: Voltage-dependent calcium channel subunit alpha-2/delta-1
type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 140 KDa

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Macromolecule #1: Voltage-dependent calcium channel subunit alpha-2/delta-1

MacromoleculeName: Voltage-dependent calcium channel subunit alpha-2/delta-1
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 124.413062 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MAAGCLLALT LTLFQSLLIG PSSEEPFPSA VTIKSWVDKM QEDLVTLAKT ASGVNQLVDI YEKYQDLYTV EPNNARQLVE IAARDIEKL LSNRSKALVR LALEAEKVQA AHQWREDFAS NEVVYYNAKD DLDPEKNDSE PGSQRIKPVF IEDANFGRQI S YQHAAVHI ...String:
MAAGCLLALT LTLFQSLLIG PSSEEPFPSA VTIKSWVDKM QEDLVTLAKT ASGVNQLVDI YEKYQDLYTV EPNNARQLVE IAARDIEKL LSNRSKALVR LALEAEKVQA AHQWREDFAS NEVVYYNAKD DLDPEKNDSE PGSQRIKPVF IEDANFGRQI S YQHAAVHI PTDIYEGSTI VLNELNWTSA LDEVFKKNRE EDPSLLWQVF GSATGLARYY PASPWVDNSR TPNKIDLYDV RR RPWYIQG AASPKDMLIL VDVSGSVSGL TLKLIRTSVS EMLETLSDDD FVNVASFNSN AQDVSCFQHL VQANVRNKKV LKD AVNNIT AKGITDYKKG FSFAFEQLLN YNVSRANCNK IIMLFTDGGE ERAQEIFNKY NKDKKVRVFT FSVGQHNYDR GPIQ WMACE NKGYYYEIPS IGAIRINTQE YLDVLGRPMV LAGDKAKQVQ WTNVYLDALE LGLVITGTLP VFNITGQFEN KTNLK NQLI LGVMGVDVSL EDIKRLTPRF TLCPNGYYFA IDPNGYVLLH PNLQPKNPKS QEPVTLDFLD AELENDIKVE IRNKMI DGE SGEKTFRTLV KSQDERYIDK GNRTYTWTPV NGTDYSLALV LPTYSFYYIK AKLEETITQA RSKKGKMKDS ETLKPDN FE ESGYTFIAPR DYCNDLKISD NNTEFLLNFN EFIDRHHHHH HHHKTPNNPS CNADLINRVL LDAGFTNELV QNYWSKQK N IKGVKARFVV TDGGITRVYP KEAGENWQEN PETYEDSFYK RSLDNDNYVF TAPYFNKSGP GAYESGIMVS KAVEIYIQG KLLKPAVVGI KIDVNSWIEN FTKTSIRDPC AGPVCDCKRN SDVMDCVILD DGGFLLMANH DDYTNQIGRF FGEIDPSLMR HLVNISVYA FNKSYDYQSV CEPGAAPKQG AGHRSAYVPS VADILQIGWW ATAAAWSILQ QFLLSLTFPR LLEAVEMEDD D FTASLSKQ SCITEQTQYF FDNDSKSFSG VLDCGNCSRI FHGEKLMNTN LIFIMVESKG TCPCDTRLLI QAEQTSDGPN PC DMVKQPR YRKGPDVCFD NNVLEDYTDC GGVSGLNPSL WYIIGIQFLL LWLVSGSTHR LL

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Macromolecule #3: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 3 / Number of copies: 7 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Macromolecule #4: 2-[(1R,5S,6S)-6-(aminomethyl)-3-ethyl-6-bicyclo[3.2.0]hept-3-enyl...

MacromoleculeName: 2-[(1R,5S,6S)-6-(aminomethyl)-3-ethyl-6-bicyclo[3.2.0]hept-3-enyl]acetic acid
type: ligand / ID: 4 / Number of copies: 1 / Formula: 8X9
Molecular weightTheoretical: 209.285 Da
Chemical component information

ChemComp-8X9:
2-[(1R,5S,6S)-6-(aminomethyl)-3-ethyl-6-bicyclo[3.2.0]hept-3-enyl]acetic acid

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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

BufferpH: 7.5
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 68.3 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6jpa

Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.23 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 207006
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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