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- PDB-8qgx: Complex between human neutrophil elastase with nanobody NbE201 -

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Basic information

Entry
Database: PDB / ID: 8qgx
TitleComplex between human neutrophil elastase with nanobody NbE201
Components
  • NbE201
  • Neutrophil elastase
KeywordsHYDROLASE / nanobody / elastase / inhibitor / serine protease
Function / homology
Function and homology information


leukocyte elastase / biosynthetic process of antibacterial peptides active against Gram-negative bacteria / neutrophil-mediated killing of fungus / negative regulation of chemotaxis / acute inflammatory response to antigenic stimulus / positive regulation of leukocyte tethering or rolling / response to yeast / leukocyte migration involved in inflammatory response / negative regulation of interleukin-8 production / negative regulation of chemokine production ...leukocyte elastase / biosynthetic process of antibacterial peptides active against Gram-negative bacteria / neutrophil-mediated killing of fungus / negative regulation of chemotaxis / acute inflammatory response to antigenic stimulus / positive regulation of leukocyte tethering or rolling / response to yeast / leukocyte migration involved in inflammatory response / negative regulation of interleukin-8 production / negative regulation of chemokine production / Antimicrobial peptides / pyroptotic inflammatory response / Activation of Matrix Metalloproteinases / neutrophil-mediated killing of gram-negative bacterium / cytokine binding / Collagen degradation / extracellular matrix disassembly / Pyroptosis / phagocytosis / response to UV / phagocytic vesicle / Degradation of the extracellular matrix / transcription repressor complex / positive regulation of MAP kinase activity / secretory granule / Regulation of Complement cascade / positive regulation of interleukin-8 production / positive regulation of smooth muscle cell proliferation / protein catabolic process / positive regulation of immune response / negative regulation of inflammatory response / intracellular calcium ion homeostasis / specific granule lumen / transcription corepressor activity / azurophil granule lumen / peptidase activity / heparin binding / protease binding / collagen-containing extracellular matrix / endopeptidase activity / response to lipopolysaccharide / defense response to bacterium / serine-type endopeptidase activity / Neutrophil degranulation / negative regulation of transcription by RNA polymerase II / cell surface / proteolysis / extracellular space / extracellular exosome / extracellular region / cytosol / cytoplasm
Similarity search - Function
: / Serine proteases, trypsin family, histidine active site / Serine proteases, trypsin family, serine active site / Serine proteases, trypsin family, histidine active site. / Peptidase S1A, chymotrypsin family / Serine proteases, trypsin family, serine active site. / Serine proteases, trypsin domain profile. / Trypsin-like serine protease / Serine proteases, trypsin domain / Trypsin ...: / Serine proteases, trypsin family, histidine active site / Serine proteases, trypsin family, serine active site / Serine proteases, trypsin family, histidine active site. / Peptidase S1A, chymotrypsin family / Serine proteases, trypsin family, serine active site. / Serine proteases, trypsin domain profile. / Trypsin-like serine protease / Serine proteases, trypsin domain / Trypsin / Peptidase S1, PA clan, chymotrypsin-like fold / Peptidase S1, PA clan
Similarity search - Domain/homology
PHOSPHATE ION / Neutrophil elastase
Similarity search - Component
Biological speciesLama glama (llama)
Homo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.3 Å
AuthorsMorales-Yanez, F. / Redeghieri, P. / Moray, J. / Dumoulin, M. / Kerff, F.
Funding support Belgium, 1items
OrganizationGrant numberCountry
Service public de Wallonie1710173 Belgium
CitationJournal: Protein Sci. / Year: 2024
Title: Enzymatic, structural, and biophysical characterization of a single-domain antibody (VHH) selectively and tightly inhibiting neutrophil elastase and exhibiting favorable developability properties.
Authors: Redeghieri, P. / Moray, J. / Kerff, F. / Gohy, S. / Leal, T. / Muyldermans, S. / Vanbever, R. / Morales-Yanez, F.J. / Dumoulin, M.
History
DepositionSep 6, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Dec 25, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Neutrophil elastase
B: NbE201
hetero molecules


Theoretical massNumber of molelcules
Total (without water)38,9065
Polymers37,6702
Non-polymers1,2363
Water32418
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: light scattering, Bio-layer interferometry
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area3850 Å2
ΔGint8 kcal/mol
Surface area14860 Å2
MethodPISA
Unit cell
Length a, b, c (Å)64.980, 73.000, 84.920
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number19
Space group name H-MP212121

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Components

#1: Protein Neutrophil elastase


Mass: 23318.982 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Homo sapiens (human) / Tissue: sputum / References: UniProt: P08246
#2: Antibody NbE201


Mass: 14351.014 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lama glama (llama) / Plasmid: pHEN6 / Production host: Escherichia coli (E. coli) / Strain (production host): WK6
#3: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta- ...2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 570.542 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4[LFucpa1-6]DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/2,3,2/[a2122h-1b_1-5_2*NCC/3=O][a1221m-1a_1-5]/1-1-2/a4-b1_a6-c1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}[(6+1)][a-L-Fucp]{}}LINUCSPDB-CARE
#4: Chemical ChemComp-PO4 / PHOSPHATE ION


Mass: 94.971 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: PO4
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 18 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestN
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.67 Å3/Da / Density % sol: 53.99 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop
Details: The crystals were grown at 20oC using the sitting drop vapor diffusion method. The drop contained 0.2 uL of hNE in complex with NbE201 at a concentration of 15 mg/mL and 0.2 uL of 0.1M HEPES ...Details: The crystals were grown at 20oC using the sitting drop vapor diffusion method. The drop contained 0.2 uL of hNE in complex with NbE201 at a concentration of 15 mg/mL and 0.2 uL of 0.1M HEPES pH7 buffer with 10 per cent (v/v) polyethylene glycol 5000 monomethyl ether and 5 per cent (v/v) tacsimate. The crystals were transferred to a cryoprotectant solution containing 33 per cent (v/v) polyethylene glycol 6000 and 33 per cent (v/v) glycerol before being frozen in liquid nitrogen.

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Data collection

DiffractionMean temperature: 293 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SOLEIL / Beamline: PROXIMA 1 / Wavelength: 0.9786 Å
DetectorType: DECTRIS EIGER X 16M / Detector: PIXEL / Date: Feb 12, 2022
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9786 Å / Relative weight: 1
ReflectionResolution: 2.3→48.54 Å / Num. obs: 18530 / % possible obs: 99.7 % / Redundancy: 13.2 % / CC1/2: 0.999 / Rmerge(I) obs: 0.128 / Net I/σ(I): 11.6
Reflection shellResolution: 2.3→2.42 Å / Rmerge(I) obs: 1.914 / Num. unique obs: 2962 / CC1/2: 0.886

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Processing

Software
NameVersionClassification
BUSTER2.10.3refinement
XDSdata reduction
XDSdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.3→48.54 Å / Cor.coef. Fo:Fc: 0.944 / Cor.coef. Fo:Fc free: 0.908 / Cross valid method: THROUGHOUT / σ(F): 0 / SU R Blow DPI: 0.297 / SU Rfree Blow DPI: 0.233
RfactorNum. reflection% reflectionSelection details
Rfree0.271 927 5 %RANDOM
Rwork0.224 ---
obs0.226 18530 99.9 %-
Displacement parametersBiso mean: 84.99 Å2
Baniso -1Baniso -2Baniso -3
1--3.9353 Å20 Å20 Å2
2---6.9655 Å20 Å2
3---10.9008 Å2
Refine analyzeLuzzati coordinate error obs: 0.43 Å
Refinement stepCycle: 1 / Resolution: 2.3→48.54 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2566 0 81 18 2665
Refine LS restraints
Refine-IDTypeDev idealNumberRestraint functionWeight
X-RAY DIFFRACTIONt_bond_d0.015310HARMONIC2
X-RAY DIFFRACTIONt_angle_deg1.139557HARMONIC2
X-RAY DIFFRACTIONt_dihedral_angle_d1177SINUSOIDAL2
X-RAY DIFFRACTIONt_incorr_chiral_ct
X-RAY DIFFRACTIONt_pseud_angle
X-RAY DIFFRACTIONt_trig_c_planes
X-RAY DIFFRACTIONt_gen_planes863HARMONIC5
X-RAY DIFFRACTIONt_it5310HARMONIC20
X-RAY DIFFRACTIONt_nbd
X-RAY DIFFRACTIONt_omega_torsion3.62
X-RAY DIFFRACTIONt_other_torsion15.78
X-RAY DIFFRACTIONt_improper_torsion
X-RAY DIFFRACTIONt_chiral_improper_torsion361SEMIHARMONIC5
X-RAY DIFFRACTIONt_sum_occupancies
X-RAY DIFFRACTIONt_utility_distance
X-RAY DIFFRACTIONt_utility_angle
X-RAY DIFFRACTIONt_utility_torsion
X-RAY DIFFRACTIONt_ideal_dist_contact5798SEMIHARMONIC4
LS refinement shellResolution: 2.3→2.44 Å / Total num. of bins used: 9
RfactorNum. reflection% reflection
Rfree0.327 148 5 %
Rwork0.2596 2814 -
all0.2628 2962 -
obs--99.7 %
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
10.640.7313-0.14220.57840.30321.9687-0.08150.02340.0856-0.14030.124-0.07010.33980.1194-0.04250.86290.0570.0203-0.0483-0.0156-0.2527-8.86461.6342-22.8571
20.2174-1.0813-0.70680.89131.91980.2859-0.0473-0.12330.0193-0.03080.0516-0.02750.0225-0.1344-0.00430.91190.06290.0041-0.05020.0008-0.2506-25.481123.7002-10.4758
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection details
1X-RAY DIFFRACTION1{ A|* }
2X-RAY DIFFRACTION2{ B|* }

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