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

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Basic information

Entry
Database: PDB / ID: 7lf1
TitleTrimeric human Arginase 1 in complex with mAb3
Components
  • Arginase-1
  • mAb3 heavy chain
  • mAb3 light chain
KeywordsHYDROLASE/IMMUNE SYSTEM / Arginase / 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 / 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)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.04 Å
AuthorsGomez-Llorente, Y. / Scapin, G. / Palte, R.L.
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, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 1, 2021Provider: repository / Type: Initial release

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Structure visualization

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Structure viewerMolecule:
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Assembly

Deposited unit
A: Arginase-1
D: mAb3 heavy chain
E: mAb3 light chain
F: mAb3 heavy chain
G: mAb3 heavy chain
H: mAb3 light chain
I: mAb3 light chain
K: mAb3 heavy chain
L: mAb3 light chain
O: mAb3 heavy chain
P: mAb3 heavy chain
Q: mAb3 light chain
R: mAb3 light chain
B: Arginase-1
C: Arginase-1
J: Arginase-1
M: Arginase-1
N: Arginase-1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)648,26130
Polymers647,60218
Non-polymers65912
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area55820 Å2
ΔGint-301 kcal/mol
Surface area167050 Å2

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Components

#1: Protein
Arginase-1 / / Liver-type arginase / Type I arginase


Mass: 34779.879 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ARG1 / Production host: Escherichia coli (E. coli) / Strain (production host): 'BL21-Gold(DE3)pLysS AG' / References: UniProt: P05089, arginase
#2: Antibody
mAb3 heavy chain


Mass: 49833.812 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): ExpiCHO-S / Production host: Cricetulus griseus (Chinese hamster)
#3: Antibody
mAb3 light chain


Mass: 23319.938 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Cell line (production host): ExpiCHO-S cells / Production host: Cricetulus griseus (Chinese hamster)
#4: Chemical
ChemComp-MN / MANGANESE (II) ION


Mass: 54.938 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: Mn
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeDetailsEntity IDParent-IDSource
1Trimeric human arginase in complex with mAb3COMPLEX2 trimers of human arginase bound to the Fab regions of 3 mAb3 molecules#1-#30MULTIPLE SOURCES
2Trimeric human arginaseCOMPLEX#11RECOMBINANT
3mAb3COMPLEX#2-#31RECOMBINANT
Molecular weightValue: 0.427 MDa / Experimental value: YES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Homo sapiens (human)9606
23Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDStrainCell
12Escherichia coli (E. coli)866768'BL21-Gold(DE3)pLysS AG'
23Cricetulus griseus (Chinese hamster)10029ExpiCHO-S
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 6 sec. / Electron dose: 44.5 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 1429
Image scansWidth: 3838 / Height: 3710

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Processing

SoftwareName: PHENIX / Version: 1.12_2829: / Classification: refinement
EM software
IDNameVersionCategory
2Leginonimage acquisition
4cryoSPARC2.0.23CTF correction
7Cootmodel fitting
9PHENIXmodel refinement
10cryoSPARC2.0.23initial Euler assignment
11cryoSPARC2.0.23final Euler assignment
12cryoSPARC2.0.23classification
13cryoSPARC2.0.233D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4.04 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 52241 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00635298
ELECTRON MICROSCOPYf_angle_d1.04647988
ELECTRON MICROSCOPYf_dihedral_angle_d6.0221108
ELECTRON MICROSCOPYf_chiral_restr0.0555454
ELECTRON MICROSCOPYf_plane_restr0.0096144

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