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- EMDB-47831: Structure of nanobody AT209 in complex with the olmesartan-bound ... -

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Basic information

Entry
Database: EMDB / ID: EMD-47831
TitleStructure of nanobody AT209 in complex with the olmesartan-bound angiotensin II type I receptor (AT1R)
Map dataLocal map of AT209-AT1RBril-olmesartan complex
Sample
  • Complex: nanobody AT209 in complex with the olmesartan-bound angiotensin II type I receptor (AT1R)
    • Protein or peptide: Nanobody AT209,Type-1 angiotensin II receptor,Soluble cytochrome b562
    • Protein or peptide: BAG2 Anti-BRIL Fab Heavy Chain
    • Protein or peptide: BAG2 Anti-BRIL Fab Light Chain
  • Ligand: Olmesartan
  • Ligand: CHOLESTEROL
KeywordsGPCR / AT1R / nanobody / MEMBRANE PROTEIN
Function / homology
Function and homology information


angiotensin type I receptor activity / angiotensin type II receptor activity / phospholipase C-activating angiotensin-activated signaling pathway / regulation of renal sodium excretion / maintenance of blood vessel diameter homeostasis by renin-angiotensin / bradykinin receptor binding / renin-angiotensin regulation of aldosterone production / positive regulation of blood vessel endothelial cell proliferation involved in sprouting angiogenesis / positive regulation of cholesterol metabolic process / low-density lipoprotein particle remodeling ...angiotensin type I receptor activity / angiotensin type II receptor activity / phospholipase C-activating angiotensin-activated signaling pathway / regulation of renal sodium excretion / maintenance of blood vessel diameter homeostasis by renin-angiotensin / bradykinin receptor binding / renin-angiotensin regulation of aldosterone production / positive regulation of blood vessel endothelial cell proliferation involved in sprouting angiogenesis / positive regulation of cholesterol metabolic process / low-density lipoprotein particle remodeling / positive regulation of macrophage derived foam cell differentiation / regulation of systemic arterial blood pressure by renin-angiotensin / regulation of vasoconstriction / Rho protein signal transduction / Peptide ligand-binding receptors / blood vessel diameter maintenance / angiotensin-activated signaling pathway / cell chemotaxis / kidney development / regulation of cell growth / calcium-mediated signaling / electron transport chain / positive regulation of reactive oxygen species metabolic process / positive regulation of inflammatory response / Cargo recognition for clathrin-mediated endocytosis / regulation of cell population proliferation / Clathrin-mediated endocytosis / positive regulation of cytosolic calcium ion concentration / regulation of inflammatory response / phospholipase C-activating G protein-coupled receptor signaling pathway / G alpha (q) signalling events / periplasmic space / electron transfer activity / G protein-coupled receptor signaling pathway / iron ion binding / inflammatory response / protein heterodimerization activity / heme binding / symbiont entry into host cell / membrane / plasma membrane
Similarity search - Function
Angiotensin II receptor type 1 / Angiotensin II receptor family / : / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562 / Serpentine type 7TM GPCR chemoreceptor Srsx / G-protein coupled receptors family 1 signature. / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM ...Angiotensin II receptor type 1 / Angiotensin II receptor family / : / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562 / Serpentine type 7TM GPCR chemoreceptor Srsx / G-protein coupled receptors family 1 signature. / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / 7 transmembrane receptor (rhodopsin family)
Similarity search - Domain/homology
Soluble cytochrome b562 / Type-1 angiotensin II receptor
Similarity search - Component
Biological speciesHomo sapiens (human) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsSkiba MA / Kruse AC
Funding support United States, 4 items
OrganizationGrant numberCountry
Helen Hay Whitney Foundation United States
National Institutes of Health/Eunice Kennedy Shriver National Institute of Child Health & Human Development (NIH/NICHD)K99HD110612 United States
National Institutes of Health/Eunice Kennedy Shriver National Institute of Child Health & Human Development (NIH/NICHD)R21HD101596 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)R01CA260415 United States
CitationJournal: Proc Natl Acad Sci U S A / Year: 2025
Title: Epitope-directed selection of GPCR nanobody ligands with evolvable function.
Authors: Meredith A Skiba / Clare Canavan / Genevieve R Nemeth / Jinghan Liu / Ali Kanso / Andrew C Kruse /
Abstract: Antibodies have the potential to target G protein-coupled receptors (GPCRs) with high receptor, cellular, and tissue selectivity; however, few antibody ligands for GPCRs exist. Here, we describe a ...Antibodies have the potential to target G protein-coupled receptors (GPCRs) with high receptor, cellular, and tissue selectivity; however, few antibody ligands for GPCRs exist. Here, we describe a generalizable selection method to enrich for GPCR ligands from a synthetic camelid antibody fragment (nanobody) library. Our strategy yielded multiple nanobody ligands for the angiotensin II type I receptor (AT1R), a prototypical GPCR and important drug target. We found that nanobodies readily act as allosteric modulators, encoding selectivity for both the receptor and chemical features of GPCR ligands. We then used structure-guided design to convert two nanobodies from allosteric ligands to competitive AT1R inhibitors through simple mutations. This work demonstrates that nanobodies can encode multiple pharmacological behaviors and have great potential as evolvable scaffolds for the development of next-generation GPCR therapeutics.
History
DepositionNov 11, 2024-
Header (metadata) releaseMar 5, 2025-
Map releaseMar 5, 2025-
UpdateMar 26, 2025-
Current statusMar 26, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_47831.png

Downloads & links

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Map

FileDownload / File: emd_47831.map.gz / Format: CCP4 / Size: 166.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationLocal map of AT209-AT1RBril-olmesartan complex
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 352 pix.
= 292.16 Å		0.83 Å/pix.
x 352 pix.
= 292.16 Å		0.83 Å/pix.
x 352 pix.
= 292.16 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.075
Minimum - Maximum-0.72860605 - 0.98600143
Average (Standard dev.)0.00011799636 (±0.010631712)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions352352352
Spacing352352352
CellA=B=C: 292.16 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Global map of AT209-AT1RBril-olmesartan complex

Fileemd_47831_additional_1.map
AnnotationGlobal map of AT209-AT1RBril-olmesartan complex
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: #2

Fileemd_47831_additional_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram (log scale)

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Additional map: #1

Fileemd_47831_additional_3.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Local map of AT209-AT1RBril-olmesartan complex, half map

Fileemd_47831_half_map_1.map
AnnotationLocal map of AT209-AT1RBril-olmesartan complex, half map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Local map of AT209-AT1RBril-olmesartan complex, half map

Fileemd_47831_half_map_2.map
AnnotationLocal map of AT209-AT1RBril-olmesartan complex, half map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : nanobody AT209 in complex with the olmesartan-bound angiotensin I...

EntireName: nanobody AT209 in complex with the olmesartan-bound angiotensin II type I receptor (AT1R)
Components
  • Complex: nanobody AT209 in complex with the olmesartan-bound angiotensin II type I receptor (AT1R)
    • Protein or peptide: Nanobody AT209,Type-1 angiotensin II receptor,Soluble cytochrome b562
    • Protein or peptide: BAG2 Anti-BRIL Fab Heavy Chain
    • Protein or peptide: BAG2 Anti-BRIL Fab Light Chain
  • Ligand: Olmesartan
  • Ligand: CHOLESTEROL

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Supramolecule #1: nanobody AT209 in complex with the olmesartan-bound angiotensin I...

SupramoleculeName: nanobody AT209 in complex with the olmesartan-bound angiotensin II type I receptor (AT1R)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Nanobody AT209,Type-1 angiotensin II receptor,Soluble cytochrome b562

MacromoleculeName: Nanobody AT209,Type-1 angiotensin II receptor,Soluble cytochrome b562
type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 65.181449 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: QVQLQESGGG LVQAGGSLRL SCAASGYIFY RYSMGWYRQA PGKEREFVAG ISYGATTNYA DSVKGRFTIS RDNAKNTVYL QMNSLKPED TAVYYCAAWG GRTLYFVYWG QGTQVTVSSG GSGEDQVDPR LIDGKILNSS TEDGIKRIQD DCPKAGRHNY I FVMIPTLY ...String:
QVQLQESGGG LVQAGGSLRL SCAASGYIFY RYSMGWYRQA PGKEREFVAG ISYGATTNYA DSVKGRFTIS RDNAKNTVYL QMNSLKPED TAVYYCAAWG GRTLYFVYWG QGTQVTVSSG GSGEDQVDPR LIDGKILNSS TEDGIKRIQD DCPKAGRHNY I FVMIPTLY SIIFVVGIFG NSLVVIVIYF YMKLKTVASV FLLNLALADL CFLLTLPLWA VYTAMEYRWP FGNYLCKIAS AS VSFNLYA SVFLLTCLSI DRYLAIVHPM KSRLRRTMLV AKVTCIIIWL LAGLASLPAI IHRNVFFIEN TNITVCAFHY ESQ NSTLPI GLGLTKNILG FLFPFLIILT SYTLIWKALK KAYDLEDNWE TLNDNLKVIE KADNAAQVKD ALTKMRAAAL DAQK ATPPK LEDKSPDSPE MKDFRHGFDI LVGQIDDALK LANEGKVKEA QAAAEQLKTT RNAYIQKYLE RARSTLDKLN DDIFK IIMA IVLFFFFSWI PHQIFTFLDV LIQLGIIRDC RIADIVDTAM PITICIAYFN NCLNPLFYGF LGKKFKRYFL QLLKYG GSS LEVLFQGPTE TSQVAPA

UniProtKB: Type-1 angiotensin II receptor, Soluble cytochrome b562, Type-1 angiotensin II receptor

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Macromolecule #2: BAG2 Anti-BRIL Fab Heavy Chain

MacromoleculeName: BAG2 Anti-BRIL Fab Heavy Chain / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 24.539314 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: EISEVQLVES GGGLVQPGGS LRLSCAASGF NVVDFSLHWV RQAPGKGLEW VAYISSSSGS TSYADSVKGR FTISADTSKN TAYLQMNSL RAEDTAVYYC ARWGYWPGEP WWKAFDYWGQ GTLVTVSSAS TKGPSVFPLA PSSKSTSGGT AALGCLVKDY F PEPVTVSW ...String:
EISEVQLVES GGGLVQPGGS LRLSCAASGF NVVDFSLHWV RQAPGKGLEW VAYISSSSGS TSYADSVKGR FTISADTSKN TAYLQMNSL RAEDTAVYYC ARWGYWPGEP WWKAFDYWGQ GTLVTVSSAS TKGPSVFPLA PSSKSTSGGT AALGCLVKDY F PEPVTVSW NSGALTSGVH TFPAVLQSSG LYSLSSVVTV PSSSLGTQTY ICNVNHKPSN TKVDKKVEPK SCD

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Macromolecule #3: BAG2 Anti-BRIL Fab Light Chain

MacromoleculeName: BAG2 Anti-BRIL Fab Light Chain / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 23.541164 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DIQMTQSPSS LSASVGDRVT ITCRASQSVS SAVAWYQQKP GKAPKLLIYS ASSLYSGVPS RFSGSRSGTD FTLTISSLQP EDFATYYCQ QYLYYSLVTF GQGTKVEIKR TVAAPSVFIF PPSDEQLKSG TASVVCLLNN FYPREAKVQW KVDNALQSGN S QESVTEQD ...String:
DIQMTQSPSS LSASVGDRVT ITCRASQSVS SAVAWYQQKP GKAPKLLIYS ASSLYSGVPS RFSGSRSGTD FTLTISSLQP EDFATYYCQ QYLYYSLVTF GQGTKVEIKR TVAAPSVFIF PPSDEQLKSG TASVVCLLNN FYPREAKVQW KVDNALQSGN S QESVTEQD SKDSTYSLSS TLTLSKADYE KHKVYACEVT HQGLSSPVTK SFNRGEC

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Macromolecule #4: Olmesartan

MacromoleculeName: Olmesartan / type: ligand / ID: 4 / Number of copies: 1 / Formula: OLM
Molecular weightTheoretical: 446.502 Da
Chemical component information

ChemComp-OLM:
Olmesartan

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Macromolecule #5: CHOLESTEROL

MacromoleculeName: CHOLESTEROL / type: ligand / ID: 5 / Number of copies: 1 / Formula: CLR
Molecular weightTheoretical: 386.654 Da
Chemical component information

ChemComp-CLR:
CHOLESTEROL

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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
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 64.1 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.8 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 903168
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 251356
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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