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- EMDB-43129: CryoEM structure of insect gustatory receptor BmGr9 -

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Basic information

Entry
Database: EMDB / ID: EMD-43129
TitleCryoEM structure of insect gustatory receptor BmGr9
Map dataMap of unbound BmGr9
Sample
  • Complex: Agonist-free BmGr9 homotetramer
    • Protein or peptide: Gustatory receptor
  • Ligand: (7R,17E,20E)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSA-17,20-DIEN-1-AMINIUM 4-OXIDE
  • Ligand: (7S)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSAN-1-AMINIUM 4-OXIDE
  • Ligand: (3beta,14beta,17beta,25R)-3-[4-methoxy-3-(methoxymethyl)butoxy]spirost-5-en
Keywordsgustatory receptor / homotetramer / ligand-gated ion channel / seven transmembrane / MEMBRANE PROTEIN
Function / homology7TM chemoreceptor / 7tm Chemosensory receptor / sensory perception of taste / signal transduction / plasma membrane / Gustatory receptor
Function and homology information
Biological speciesBombyx mori (domestic silkworm)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.85 Å
AuthorsFrank HM / Walsh Jr RM / Garrity PA / Gaudet R
Funding support United States, 3 items
OrganizationGrant numberCountry
Other governmentR21DC018497
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM120996 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01AI157194 United States
CitationJournal: bioRxiv / Year: 2023
Title: Structure of an insect gustatory receptor.
Authors: Heather M Frank / Sanket Walujkar / Richard M Walsh / Willem J Laursen / Douglas L Theobald / Paul A Garrity / Rachelle Gaudet /
Abstract: Gustatory Receptors (GRs) are critical for insect chemosensation and are potential targets for controlling pests and disease vectors. However, GR structures have not been experimentally determined. ...Gustatory Receptors (GRs) are critical for insect chemosensation and are potential targets for controlling pests and disease vectors. However, GR structures have not been experimentally determined. We present structures of Gr9 (BmGr9), a fructose-gated cation channel, in agonist-free and fructose-bound states. BmGr9 forms a tetramer similar to distantly related insect Olfactory Receptors (ORs). Upon fructose binding, BmGr9's ion channel gate opens through helix S7b movements. In contrast to ORs, BmGR9's ligand-binding pocket, shaped by a kinked helix S4 and a shorter extracellular S3-S4 loop, is larger and solvent accessible in both agonist-free and fructose-bound states. Also unlike ORs, fructose binding by BmGr9 involves helix S5 and a binding pocket lined with aromatic and polar residues. Structure-based sequence alignments reveal distinct patterns of ligand-binding pocket residue conservation in GR subfamilies associated with distinct ligand classes. These data provide insight into the molecular basis of GR ligand specificity and function.
History
DepositionDec 13, 2023-
Header (metadata) releaseJan 10, 2024-
Map releaseJan 10, 2024-
UpdateJan 24, 2024-
Current statusJan 24, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43129.png

Downloads & links

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Map

FileDownload / File: emd_43129.map.gz / Format: CCP4 / Size: 93 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMap of unbound BmGr9
Voxel sizeX=Y=Z: 0.83 Å
Density
Contour LevelBy AUTHOR: 0.186
Minimum - Maximum-1.9138396 - 2.9011216
Average (Standard dev.)0.002026128 (±0.06676248)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions290290290
Spacing290290290
CellA=B=C: 240.7 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half map A of unbound BmGr9

Fileemd_43129_half_map_1.map
AnnotationHalf map A of unbound BmGr9
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B of unbound BmGr9

Fileemd_43129_half_map_2.map
AnnotationHalf map B of unbound BmGr9
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Agonist-free BmGr9 homotetramer

EntireName: Agonist-free BmGr9 homotetramer
Components
  • Complex: Agonist-free BmGr9 homotetramer
    • Protein or peptide: Gustatory receptor
  • Ligand: (7R,17E,20E)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSA-17,20-DIEN-1-AMINIUM 4-OXIDE
  • Ligand: (7S)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSAN-1-AMINIUM 4-OXIDE
  • Ligand: (3beta,14beta,17beta,25R)-3-[4-methoxy-3-(methoxymethyl)butoxy]spirost-5-en

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Supramolecule #1: Agonist-free BmGr9 homotetramer

SupramoleculeName: Agonist-free BmGr9 homotetramer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Bombyx mori (domestic silkworm)
Molecular weightTheoretical: 218.05 KDa

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Macromolecule #1: Gustatory receptor

MacromoleculeName: Gustatory receptor / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Bombyx mori (domestic silkworm)
Molecular weightTheoretical: 54.57232 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGWSHPQFEK GGGSGGGSGG SAWSHPQFEK AALEVLFQGP GTMPPSPDLR ADEPKTPCLV GGAHAFILKI SSFCGLAPLR FEPRSQEYA VTISKGKCFY SYILVTFLVI CTIYGLVAEI GVGVEKSVRM SSRMSQVVSA CDILVVAVTA GVGVYGAPAR M RTMLSYME ...String:
MGWSHPQFEK GGGSGGGSGG SAWSHPQFEK AALEVLFQGP GTMPPSPDLR ADEPKTPCLV GGAHAFILKI SSFCGLAPLR FEPRSQEYA VTISKGKCFY SYILVTFLVI CTIYGLVAEI GVGVEKSVRM SSRMSQVVSA CDILVVAVTA GVGVYGAPAR M RTMLSYME NIVAVDRELG RHHSAATERK LCALLLLILL SFTILLVDDF CFYAMQAGKT GRQWEIVTNY AGFYFLWYIV MV LELQFAF TALSLRARLK LFNEALNVTA SQVCKPVKKP KNSQLSVYAT SVRPVSCKRE NVIVETIRVR DKDDAFVMMK TAD GVPCLQ VPPCEAVGRL SRMRCTLCEV TRHIADGYGL PLVIILMSTL LHLIVTPYFL IMEIIVSTHR LHFLVLQFLW CTTH LIRML VVVEPCHYTM REGKRTEDIV CRLMTLAPHG GVLSSKLEVL SRLLMLQNIS YSPLGMCTLD RPLIVTVLGA VTTYL VILI QFQRYDS

UniProtKB: Gustatory receptor

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Macromolecule #2: (7R,17E,20E)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)MET...

MacromoleculeName: (7R,17E,20E)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSA-17,20-DIEN-1-AMINIUM 4-OXIDE
type: ligand / ID: 2 / Number of copies: 8 / Formula: PSC
Molecular weightTheoretical: 759.068 Da
Chemical component information

ChemComp-PSC:
(7R,17E,20E)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSA-17,20-DIEN-1-AMINIUM 4-OXIDE / phospholipid*YM / Phosphatidylcholine

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Macromolecule #3: (7S)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5...

MacromoleculeName: (7S)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSAN-1-AMINIUM 4-OXIDE
type: ligand / ID: 3 / Number of copies: 20 / Formula: PC7
Molecular weightTheoretical: 763.1 Da
Chemical component information

ChemComp-PC7:
(7S)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSAN-1-AMINIUM 4-OXIDE / phospholipid*YM / Phosphatidylcholine

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Macromolecule #4: (3beta,14beta,17beta,25R)-3-[4-methoxy-3-(methoxymethyl)butoxy]sp...

MacromoleculeName: (3beta,14beta,17beta,25R)-3-[4-methoxy-3-(methoxymethyl)butoxy]spirost-5-en
type: ligand / ID: 4 / Number of copies: 4 / Formula: 9Z9
Molecular weightTheoretical: 544.805 Da
Chemical component information

ChemComp-9Z9:
(3beta,14beta,17beta,25R)-3-[4-methoxy-3-(methoxymethyl)butoxy]spirost-5-en / detergent*YM

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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: 8.25
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated magnification: 60240 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 105000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 11474 / Average exposure time: 3.803 sec. / Average electron dose: 74.492 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1955549
Startup modelType of model: INSILICO MODEL
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4.0.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final reconstructionApplied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.85 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.2) / Number images used: 1011451
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: Other / Chain - Initial model type: in silico model / Details: ColabFold generated
DetailsInitial fitting was done using DockInMap in PHENIX and refined through cycles of manual rebuilding in Coot, real-space refinement in PHENIX, and remodeling by simulations run in the ISOLDE plugin of ChimeraX.
RefinementSpace: REAL / Protocol: FLEXIBLE FIT / Overall B value: 58.19 / Target criteria: cross-correlation coefficient
Output model

PDB-8vc1:
CryoEM structure of insect gustatory receptor BmGr9

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