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- EMDB-23296: Trimeric human Arginase 1 in complex with mAb2 -

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Basic information

Entry
Database: EMDB / ID: EMD-23296
TitleTrimeric human Arginase 1 in complex with mAb2
Map dataSharpened map, focused on one hARG trimer for the complex 2x3hARG:3Mab2
Sample
  • Complex: Trimeric human arginase in complex with mAb2
    • Complex: Trimeric human arginase
      • Protein or peptide: Arginase-1
    • Complex: mAb2
      • Protein or peptide: mAb2 heavy chain
      • Protein or peptide: mAb2 light chain
KeywordsArginase / Metalloenzyme / IMMUNE SYSTEM / HYDROLASE-IMMUNE SYSTEM complex
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 / arginase / arginase activity / : / urea cycle / response to nematode / defense response to protozoan / negative regulation of type II interferon-mediated signaling pathway ...positive regulation of neutrophil mediated killing of fungus / Urea cycle / negative regulation of T-helper 2 cell cytokine production / arginase / arginase activity / : / urea cycle / response to nematode / defense response to protozoan / negative regulation of type II interferon-mediated signaling pathway / negative regulation of activated T cell proliferation / L-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: 3.68 Å
AuthorsGomez-Llorente Y / Scapin G
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-
UpdateOct 23, 2024-
Current statusOct 23, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.38
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.38
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7lf0
  • Surface level: 0.38
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23296.png

Downloads & links

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Map

FileDownload / File: emd_23296.map.gz / Format: CCP4 / Size: 147.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened map, focused on one hARG trimer for the complex 2x3hARG:3Mab2
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.04 Å/pix.
x 338 pix.
= 351.52 Å		1.04 Å/pix.
x 338 pix.
= 351.52 Å		1.04 Å/pix.
x 338 pix.
= 351.52 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.04 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.38 / Movie #1: 0.38
Minimum - Maximum-11.263761499999999 - 9.835300999999999
Average (Standard dev.)0.0052860584 (±0.08175929)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions338338338
Spacing338338338
CellA=B=C: 351.52 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.041.041.04
M x/y/z338338338
origin x/y/z0.0000.0000.000
length x/y/z351.520351.520351.520
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS338338338
D min/max/mean-11.2649.8350.005

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Supplemental data

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Mask #1

Fileemd_23296_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Filtered map at the final resolution.

Fileemd_23296_additional_1.map
AnnotationFiltered map at the final resolution.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map A.

Fileemd_23296_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_23296_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 mAb2

EntireName: Trimeric human arginase in complex with mAb2
Components
  • Complex: Trimeric human arginase in complex with mAb2
    • Complex: Trimeric human arginase
      • Protein or peptide: Arginase-1
    • Complex: mAb2
      • Protein or peptide: mAb2 heavy chain
      • Protein or peptide: mAb2 light chain

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Supramolecule #1: Trimeric human arginase in complex with mAb2

SupramoleculeName: Trimeric human arginase in complex with mAb2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: 2 trimers of human arginase bound to the Fab regions of 3 mAb2 molecules.
Molecular weightTheoretical: 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)

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Supramolecule #3: mAb2

SupramoleculeName: mAb2 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2-#3
Source (natural)Organism: Homo sapiens (human)

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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

UniProtKB: Arginase-1

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Macromolecule #2: mAb2 heavy chain

MacromoleculeName: mAb2 heavy chain / type: protein_or_peptide / ID: 2 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 49.573676 KDa
Recombinant expressionOrganism: Cricetulus griseus (Chinese hamster)
SequenceString: QVQLVQSGAE VKKPGASVKV SCKASGYTFT NYGISWVRQA PGQGLEWMGW ISAYNGNTNY AQKLQGRVTM TTDTSTSTAY MELRSLRSD DTAVYYCARE GAYGYRSPYH NWFDPWGQGT LVTVSSAKTT PPSVYPLAPG SAAQTNSMVT LGCLVKGYFP E PVTVTWNS ...String:
QVQLVQSGAE VKKPGASVKV SCKASGYTFT NYGISWVRQA PGQGLEWMGW ISAYNGNTNY AQKLQGRVTM TTDTSTSTAY MELRSLRSD DTAVYYCARE GAYGYRSPYH NWFDPWGQGT LVTVSSAKTT PPSVYPLAPG SAAQTNSMVT LGCLVKGYFP E PVTVTWNS GSLSSGVHTF PAVLQSDLYT LSSSVTVPSS TWPSETVTCN VAHPASSTKV DKKIVPRDCG CKPCICTVPE VS SVFIFPP KPKDVLTITL TPKVTCVVVA ISKDDPEVQF SWFVDDVEVH TAQTQPREEQ FNSTFRSVSE LPIMHQDWLN GKE FKCRVN SAAFPAPIEK TISKTKGRPK APQVYTIPPP KEQMAKDKVS LTCMITDFFP EDITVEWQWN GQPAENYKNT QPIM DTDGS YFVYSKLNVQ KSNWEAGNTF TCSVLHEGLH NHHTEKSLSH SPGK

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Macromolecule #3: mAb2 light chain

MacromoleculeName: mAb2 light chain / type: protein_or_peptide / ID: 3 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 23.488965 KDa
Recombinant expressionOrganism: Cricetulus griseus (Chinese hamster)
SequenceString: EIVMTQSPAT LSLSPGERAT LSCRASQSVS SYLAWYQQKP GQAPRLLIYD ASNRATGIPA RFSGSGSGTD FTLTISSLEP EDFAVYYCQ QHSLLPRTFG GGTKVEIKRA DAAPTVSIFP PSSEQLTSGG ASVVCFLNNF YPKDINVKWK IDGSERQNGV L NSWTDQDS ...String:
EIVMTQSPAT LSLSPGERAT LSCRASQSVS SYLAWYQQKP GQAPRLLIYD ASNRATGIPA RFSGSGSGTD FTLTISSLEP EDFAVYYCQ QHSLLPRTFG GGTKVEIKRA DAAPTVSIFP PSSEQLTSGG ASVVCFLNNF YPKDINVKWK IDGSERQNGV L NSWTDQDS KDSTYSMSST LTLTKDEYER HNSYTCEATH KTSTSPIVKS FNRNEC

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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: C-flat-1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 30 sec.
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
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: 1197 / Average exposure time: 6.0 sec. / Average electron dose: 44.3 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.2 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.68 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.0.23) / Number images used: 52550
Initial angle assignmentType: OTHER / Software - Name: cryoSPARC (ver. 2.0.23)
Final angle assignmentType: OTHER / Software - Name: cryoSPARC (ver. 2.0.23)
Final 3D classificationNumber classes: 3 / Software - Name: cryoSPARC (ver. 2.0.23)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-7lf0:
Trimeric human Arginase 1 in complex with mAb2

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