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- EMDB-23297: Trimeric human Arginase 1 in complex with mAb3 -

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Basic information

Entry
Database: EMDB / ID: EMD-23297
TitleTrimeric human Arginase 1 in complex with mAb3
Map dataFull map of 2 hARG trimers bound to 3 Mab3 molecules (only the Fab regions are visible), flipped.
Sample
  • Complex: Trimeric human arginase in complex with mAb3
    • Complex: Trimeric human arginase
      • Protein or peptide: Arginase-1
    • Complex: mAb3
      • Protein or peptide: mAb3 heavy chain
      • Protein or peptide: mAb3 light chain
  • Ligand: MANGANESE (II) ION
Function / homology
Function and homology information


positive regulation of neutrophil mediated killing of fungus / Urea cycle / negative regulation of T-helper 2 cell cytokine production / arginine catabolic process to ornithine / arginase / arginase activity / urea cycle / negative regulation of type II interferon-mediated signaling pathway / defense response to protozoan / negative regulation of activated T cell proliferation ...positive regulation of neutrophil mediated killing of fungus / Urea cycle / negative regulation of T-helper 2 cell cytokine production / arginine catabolic process to ornithine / arginase / arginase activity / urea cycle / negative regulation of type II interferon-mediated signaling pathway / defense response to protozoan / negative regulation of activated T cell proliferation / arginine catabolic process / negative regulation of T cell proliferation / specific granule lumen / azurophil granule lumen / manganese ion binding / adaptive immune response / innate immune response / Neutrophil degranulation / extracellular space / extracellular region / nucleus / cytosol / cytoplasm
Similarity search - Function
Arginase / Ureohydrolase, manganese-binding site / Arginase family signature. / Ureohydrolase / Arginase family / Arginase family profile. / Ureohydrolase domain superfamily
Similarity search - Domain/homology
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.04 Å
AuthorsGomez-Llorente Y / Scapin G / Palte RL
CitationJournal: Commun Biol / Year: 2021
Title: Cryo-EM structures of inhibitory antibodies complexed with arginase 1 provide insight into mechanism of action.
Authors: Rachel L Palte / Veronica Juan / Yacob Gomez-Llorente / Marc Andre Bailly / Kalyan Chakravarthy / Xun Chen / Daniel Cipriano / Ghassan N Fayad / Laurence Fayadat-Dilman / Symon Gathiaka / ...Authors: Rachel L Palte / Veronica Juan / Yacob Gomez-Llorente / Marc Andre Bailly / Kalyan Chakravarthy / Xun Chen / Daniel Cipriano / Ghassan N Fayad / Laurence Fayadat-Dilman / Symon Gathiaka / Heiko Greb / Brian Hall / Mas Handa / Mark Hsieh / Esther Kofman / Heping Lin / J Richard Miller / Nhung Nguyen / Jennifer O'Neil / Hussam Shaheen / Eric Sterner / Corey Strickland / Angie Sun / Shane Taremi / Giovanna Scapin /
Abstract: Human Arginase 1 (hArg1) is a metalloenzyme that catalyzes the hydrolysis of L-arginine to L-ornithine and urea, and modulates T-cell-mediated immune response. Arginase-targeted therapies have been ...Human Arginase 1 (hArg1) is a metalloenzyme that catalyzes the hydrolysis of L-arginine to L-ornithine and urea, and modulates T-cell-mediated immune response. Arginase-targeted therapies have been pursued across several disease areas including immunology, oncology, nervous system dysfunction, and cardiovascular dysfunction and diseases. Currently, all published hArg1 inhibitors are small molecules usually less than 350 Da in size. Here we report the cryo-electron microscopy structures of potent and inhibitory anti-hArg antibodies bound to hArg1 which form distinct macromolecular complexes that are greater than 650 kDa. With local resolutions of 3.5 Å or better we unambiguously mapped epitopes and paratopes for all five antibodies and determined that the antibodies act through orthosteric and allosteric mechanisms. These hArg1:antibody complexes present an alternative mechanism to inhibit hArg1 activity and highlight the ability to utilize antibodies as probes in the discovery and development of peptide and small molecule inhibitors for enzymes in general.
History
DepositionJan 15, 2021-
Header (metadata) releaseSep 1, 2021-
Map releaseSep 1, 2021-
UpdateSep 1, 2021-
Current statusSep 1, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.36
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.36
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7lf1
  • Surface level: 0.36
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23297.png

Downloads & links

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Map

FileDownload / File: emd_23297.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFull map of 2 hARG trimers bound to 3 Mab3 molecules (only the Fab regions are visible), flipped.
Voxel sizeX=Y=Z: 1.04 Å
Density
Contour LevelBy AUTHOR: 0.36 / Movie #1: 0.36
Minimum - Maximum-0.79983616 - 1.7726858
Average (Standard dev.)0.0098683415 (±0.0567408)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 332.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.041.041.04
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z332.800332.800332.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.8001.7730.010

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Supplemental data

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Mask #1

Fileemd_23297_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A.

Fileemd_23297_half_map_1.map
AnnotationHalf map A.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B.

Fileemd_23297_half_map_2.map
AnnotationHalf map B.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Trimeric human arginase in complex with mAb3

EntireName: Trimeric human arginase in complex with mAb3
Components
  • Complex: Trimeric human arginase in complex with mAb3
    • Complex: Trimeric human arginase
      • Protein or peptide: Arginase-1
    • Complex: mAb3
      • Protein or peptide: mAb3 heavy chain
      • Protein or peptide: mAb3 light chain
  • Ligand: MANGANESE (II) ION

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Supramolecule #1: Trimeric human arginase in complex with mAb3

SupramoleculeName: Trimeric human arginase in complex with mAb3 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Details: 2 trimers of human arginase bound to the Fab regions of 3 mAb3 molecules
Molecular weightExperimental: 427 KDa

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Supramolecule #2: Trimeric human arginase

SupramoleculeName: Trimeric human arginase / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Escherichia coli (E. coli) / Recombinant strain: 'BL21-Gold(DE3)pLysS AG'

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Supramolecule #3: mAb3

SupramoleculeName: mAb3 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2-#3
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Cricetulus griseus (Chinese hamster) / Recombinant cell: ExpiCHO-S

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Macromolecule #1: Arginase-1

MacromoleculeName: Arginase-1 / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO / EC number: arginase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 34.779879 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MSAKSRTIGI IGAPFSKGQP RGGVEEGPTV LRKAGLLEKL KEQECDVKDY GDLPFADIPN DSPFQIVKNP RSVGKASEQL AGKVAEVKK NGRISLVLGG DHSLAIGSIS GHARVHPDLG VIWVDAHTDI NTPLTTTSGN LHGQPVSFLL KELKGKIPDV P GFSWVTPC ...String:
MSAKSRTIGI IGAPFSKGQP RGGVEEGPTV LRKAGLLEKL KEQECDVKDY GDLPFADIPN DSPFQIVKNP RSVGKASEQL AGKVAEVKK NGRISLVLGG DHSLAIGSIS GHARVHPDLG VIWVDAHTDI NTPLTTTSGN LHGQPVSFLL KELKGKIPDV P GFSWVTPC ISAKDIVYIG LRDVDPGEHY ILKTLGIKYF SMTEVDRLGI GKVMEETLSY LLGRKKRPIH LSFDVDGLDP SF TPATGTP VVGGLTYREG LYITEEIYKT GLLSGLDIME VNPSLGKTPE EVTRTVNTAV AITLACFGLA REGNHKPIDY LNP PK

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Macromolecule #2: mAb3 heavy chain

MacromoleculeName: mAb3 heavy chain / type: protein_or_peptide / ID: 2 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 49.833812 KDa
Recombinant expressionOrganism: Cricetulus griseus (Chinese hamster)
SequenceString: EVQLVQSGAE VKKPGASVKV SCKASGYTFF KYGISWVRQA PGQGLEWMGS ISPYTGETHY AQKLQGRVTM TTDTSTSTAY MELRSLRSD DTAVYYCARE GAYGYRSPYQ NWFDPWGQGT LVTVSSASTK GPSVFPLAPC SRSTSESTAA LGCLVKDYFP E PVTVSWNS ...String:
EVQLVQSGAE VKKPGASVKV SCKASGYTFF KYGISWVRQA PGQGLEWMGS ISPYTGETHY AQKLQGRVTM TTDTSTSTAY MELRSLRSD DTAVYYCARE GAYGYRSPYQ NWFDPWGQGT LVTVSSASTK GPSVFPLAPC SRSTSESTAA LGCLVKDYFP E PVTVSWNS GALTSGVHTF PAVLQSSGLY SLSSVVTVPS SSLGTKTYTC NVDHKPSNTK VDKRVESKYG PPCPPCPAPE FL GGPSVFL FPPKPKDTLM ISRTPEVTCV VVDVSQEDPE VQFNWYVDGV EVHNAKTKPR EEQFNSTYRV VSVLTVLHQD WLN GKEYKC KVSNKGLPSS IEKTISKAKG QPREPQVYTL PPSQEEMTKN QVSLTCLVKG FYPSDIAVEW ESNGQPENNY KTTP PVLDS DGSFFLYSRL TVDKSRWQEG NVFSCSVMHE ALHNHYTQKS LSLSLGK

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Macromolecule #3: mAb3 light chain

MacromoleculeName: mAb3 light chain / type: protein_or_peptide / ID: 3 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 23.319938 KDa
Recombinant expressionOrganism: Cricetulus griseus (Chinese hamster)
SequenceString: EIVMTQSPAT LSLSPGERAT LSCRASQSVS SYLAWYQQKP GQAPRLLIYD ASNRATGIPA RFSGSGSGTD FTLTISSLEP EDFAVYYCQ QHSLLPRTFG GGTKVEIKRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF YPREAKVQWK VDNALQSGNS Q ESVTEQDS ...String:
EIVMTQSPAT LSLSPGERAT LSCRASQSVS SYLAWYQQKP GQAPRLLIYD ASNRATGIPA RFSGSGSGTD FTLTISSLEP EDFAVYYCQ QHSLLPRTFG GGTKVEIKRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF YPREAKVQWK VDNALQSGNS Q ESVTEQDS KDSTYSLSST LTLSKADYEK HKVYACEVTH QGLSSPVTKS FNRGEC

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Macromolecule #4: MANGANESE (II) ION

MacromoleculeName: MANGANESE (II) ION / type: ligand / ID: 4 / Number of copies: 12 / Formula: MN
Molecular weightTheoretical: 54.938 Da

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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: 7.4
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE

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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: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Number grids imaged: 1 / Number real images: 1429 / Average exposure time: 6.0 sec. / Average electron dose: 44.5 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: cryoSPARC (ver. 2.0.23)
Initial angle assignmentType: OTHER / Software - Name: cryoSPARC (ver. 2.0.23)
Final 3D classificationNumber classes: 3 / Software - Name: cryoSPARC (ver. 2.0.23)
Final angle assignmentType: OTHER / Software - Name: cryoSPARC (ver. 2.0.23)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.04 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.0.23) / Number images used: 52241
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-7lf1:
Trimeric human Arginase 1 in complex with mAb3

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