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- PDB-8q95: Crystal structure of the SARS-CoV-2 BA.1 RBD with neutralizing-VH... -

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Basic information

Entry
Database: PDB / ID: 8q95
TitleCrystal structure of the SARS-CoV-2 BA.1 RBD with neutralizing-VHHs Ma16B06 and Ma3F05
Components
  • Nanobody Ma16B06
  • Nanobody Ma3F05
  • Spike protein S1
KeywordsVIRAL PROTEIN / Neutralizing VHH / Fold-promoter
Function / homology
Function and homology information


Maturation of spike protein / viral translation / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / suppression by virus of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / entry receptor-mediated virion attachment to host cell ...Maturation of spike protein / viral translation / Translation of Structural Proteins / Virion Assembly and Release / host cell surface / host extracellular space / suppression by virus of host tetherin activity / Induction of Cell-Cell Fusion / structural constituent of virion / entry receptor-mediated virion attachment to host cell / host cell endoplasmic reticulum-Golgi intermediate compartment membrane / receptor-mediated endocytosis of virus by host cell / membrane fusion / Attachment and Entry / positive regulation of viral entry into host cell / receptor-mediated virion attachment to host cell / receptor ligand activity / host cell surface receptor binding / fusion of virus membrane with host plasma membrane / fusion of virus membrane with host endosome membrane / viral envelope / virion attachment to host cell / SARS-CoV-2 activates/modulates innate and adaptive immune responses / host cell plasma membrane / virion membrane / identical protein binding / membrane / plasma membrane
Similarity search - Function
Spike (S) protein S1 subunit, receptor-binding domain, SARS-CoV-2 / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike glycoprotein, betacoronavirus / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus / Betacoronavirus spike glycoprotein S1, receptor binding / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like ...Spike (S) protein S1 subunit, receptor-binding domain, SARS-CoV-2 / Spike (S) protein S1 subunit, N-terminal domain, SARS-CoV-like / Betacoronavirus spike (S) glycoprotein S1 subunit N-terminal (NTD) domain profile. / Spike glycoprotein, N-terminal domain superfamily / Betacoronavirus spike (S) glycoprotein S1 subunit C-terminal (CTD) domain profile. / Spike glycoprotein, betacoronavirus / Spike (S) protein S1 subunit, receptor-binding domain, betacoronavirus / Spike S1 subunit, receptor binding domain superfamily, betacoronavirus / Betacoronavirus spike glycoprotein S1, receptor binding / Spike glycoprotein S1, N-terminal domain, betacoronavirus-like / Betacoronavirus-like spike glycoprotein S1, N-terminal / Spike glycoprotein S2, coronavirus, heptad repeat 1 / Spike glycoprotein S2, coronavirus, heptad repeat 2 / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 2 (HR2) region profile. / Coronavirus spike (S) glycoprotein S2 subunit heptad repeat 1 (HR1) region profile. / Spike glycoprotein S2 superfamily, coronavirus / Spike glycoprotein S2, coronavirus / Coronavirus spike glycoprotein S2 / Coronavirus spike glycoprotein S1, C-terminal / Coronavirus spike glycoprotein S1, C-terminal
Similarity search - Domain/homology
Biological speciesSevere acute respiratory syndrome coronavirus 2
Vicugna pacos (alpaca)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.6 Å
AuthorsAksu, M. / Rymarenko, O. / Guttler, T. / Gorlich, D.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Antiviral Res / Year: 2024
Title: Nanobodies to multiple spike variants and inhalation of nanobody-containing aerosols neutralize SARS-CoV-2 in cell culture and hamsters.
Authors: Metin Aksu / Priya Kumar / Thomas Güttler / Waltraud Taxer / Kathrin Gregor / Bianka Mußil / Oleh Rymarenko / Kim M Stegmann / Antje Dickmanns / Sabrina Gerber / Wencke Reineking / Claudia ...Authors: Metin Aksu / Priya Kumar / Thomas Güttler / Waltraud Taxer / Kathrin Gregor / Bianka Mußil / Oleh Rymarenko / Kim M Stegmann / Antje Dickmanns / Sabrina Gerber / Wencke Reineking / Claudia Schulz / Timo Henneck / Ahmed Mohamed / Gerhard Pohlmann / Mehmet Ramazanoglu / Kemal Mese / Uwe Groß / Tamar Ben-Yedidia / Oded Ovadia / Dalit Weinstein Fischer / Merav Kamensky / Amir Reichman / Wolfgang Baumgärtner / Maren von Köckritz-Blickwede / Matthias Dobbelstein / Dirk Görlich /
Abstract: The ongoing threat of COVID-19 has highlighted the need for effective prophylaxis and convenient therapies, especially for outpatient settings. We have previously developed highly potent single- ...The ongoing threat of COVID-19 has highlighted the need for effective prophylaxis and convenient therapies, especially for outpatient settings. We have previously developed highly potent single-domain (VHH) antibodies, also known as nanobodies, that target the Receptor Binding Domain (RBD) of the SARS-CoV-2 Spike protein and neutralize the Wuhan strain of the virus. In this study, we present a new generation of anti-RBD nanobodies with superior properties. The primary representative of this group, Re32D03, neutralizes Alpha to Delta as well as Omicron BA.2.75; other members neutralize, in addition, Omicron BA.1, BA.2, BA.4/5, and XBB.1. Crystal structures of RBD-nanobody complexes reveal how ACE2-binding is blocked and also explain the nanobodies' tolerance to immune escape mutations. Through the cryo-EM structure of the Ma16B06-BA.1 Spike complex, we demonstrated how a single nanobody molecule can neutralize a trimeric spike. We also describe a method for large-scale production of these nanobodies in Pichia pastoris, and for formulating them into aerosols. Exposing hamsters to these aerosols, before or even 24 h after infection with SARS-CoV-2, significantly reduced virus load, weight loss and pathogenicity. These results show the potential of aerosolized nanobodies for prophylaxis and therapy of coronavirus infections.
History
DepositionAug 19, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Dec 20, 2023Provider: repository / Type: Initial release
Revision 1.1Dec 27, 2023Group: Database references / Category: citation / Item: _citation.journal_volume

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Spike protein S1
B: Nanobody Ma3F05
C: Nanobody Ma16B06


Theoretical massNumber of molelcules
Total (without water)48,8193
Polymers48,8193
Non-polymers00
Water6,395355
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area2670 Å2
ΔGint-15 kcal/mol
Surface area18900 Å2
MethodPISA
Unit cell
Length a, b, c (Å)82.222, 89.560, 72.198
Angle α, β, γ (deg.)90.00, 111.63, 90.00
Int Tables number5
Space group name H-MC121

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Components

#1: Protein Spike protein S1


Mass: 21375.975 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Severe acute respiratory syndrome coronavirus 2
Gene: S, 2 / Production host: Komagataella pastoris (fungus) / References: UniProt: P0DTC2
#2: Antibody Nanobody Ma3F05


Mass: 14260.616 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vicugna pacos (alpaca) / Production host: Komagataella pastoris (fungus)
#3: Antibody Nanobody Ma16B06


Mass: 13182.485 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vicugna pacos (alpaca) / Production host: Escherichia coli (E. coli)
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 355 / Source method: isolated from a natural source / Formula: H2O

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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: 2.53 Å3/Da / Density % sol: 51.4 %
Crystal growTemperature: 293 K / Method: vapor diffusion / pH: 7.8
Details: 0.1 M HEPES, 20% (v/v) PEG smear high, 0.15 M Lithium sulphate, 0.05 M magnesium chloride

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SLS / Beamline: X10SA / Wavelength: 1 Å
DetectorType: DECTRIS EIGER2 X 16M / Detector: PIXEL / Date: May 9, 2022
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 1.45→38.22 Å / Num. obs: 165939 / % possible obs: 97.4 % / Redundancy: 6.7 % / CC1/2: 0.99 / Rmerge(I) obs: 0.081 / Rrim(I) all: 0.096 / Net I/σ(I): 8.08
Reflection shell
Resolution (Å)Rmerge(I) obsNum. unique obsCC1/2Rrim(I) allDiffraction-ID
1.45-1.541.5261650.461.8081
1.54-1.640.756251830.7310.9081
1.64-1.770.405233870.9060.481
1.77-1.940.202216460.9650.241
1.94-2.170.112196370.980.1351
2.17-2.510.085173900.9860.11
2.51-3.070.073147630.9880.0861
3.07-4.330.062114580.990.0741
4.33-38.220.06463100.990.0761

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Processing

Software
NameVersionClassification
PHENIX1.20.1_4487refinement
XSCALEdata scaling
XDSdata reduction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 1.6→38.22 Å / SU ML: 0.18 / Cross valid method: THROUGHOUT / σ(F): 1.37 / Phase error: 21.38 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2062 3159 5 %
Rwork0.1755 --
obs0.177 63162 98.68 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.1 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 1.6→38.22 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3325 0 0 355 3680
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0153458
X-RAY DIFFRACTIONf_angle_d1.3324723
X-RAY DIFFRACTIONf_dihedral_angle_d12.7771227
X-RAY DIFFRACTIONf_chiral_restr0.089500
X-RAY DIFFRACTIONf_plane_restr0.012614
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
1.6-1.620.29071320.2592525X-RAY DIFFRACTION98
1.62-1.650.29261360.24242574X-RAY DIFFRACTION97
1.65-1.680.26971360.20132578X-RAY DIFFRACTION98
1.68-1.710.23631350.19132576X-RAY DIFFRACTION98
1.71-1.740.22091380.19142604X-RAY DIFFRACTION98
1.74-1.770.23321340.18622549X-RAY DIFFRACTION98
1.77-1.810.2451360.18852588X-RAY DIFFRACTION98
1.81-1.840.21121380.18842629X-RAY DIFFRACTION98
1.85-1.890.23731350.19312557X-RAY DIFFRACTION98
1.89-1.940.22051370.18862590X-RAY DIFFRACTION98
1.94-1.990.23451370.18092608X-RAY DIFFRACTION99
1.99-2.050.21681380.1772617X-RAY DIFFRACTION99
2.05-2.110.22631370.17882610X-RAY DIFFRACTION99
2.11-2.190.18061380.17992624X-RAY DIFFRACTION99
2.19-2.270.21941370.1782604X-RAY DIFFRACTION99
2.28-2.380.21341360.18612588X-RAY DIFFRACTION99
2.38-2.50.21811400.1822660X-RAY DIFFRACTION99
2.5-2.660.23711390.18572625X-RAY DIFFRACTION99
2.66-2.870.21231380.1792633X-RAY DIFFRACTION100
2.87-3.150.20921400.18042652X-RAY DIFFRACTION100
3.15-3.610.19771390.16612652X-RAY DIFFRACTION100
3.61-4.550.18861400.14812659X-RAY DIFFRACTION100
4.55-38.220.16211430.16672701X-RAY DIFFRACTION100
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)
15.7155-4.53642.04957.9603-2.08562.3260.16560.21340.2447-0.3282-0.2580.3063-0.0315-0.00110.11210.18960.07220.01650.2599-0.03130.2661-4.098-10.05818.342
27.7924-0.15711.53645.46430.55938.4296-0.1632-0.33360.30470.11740.1152-0.36990.00120.17040.03780.14370.0593-0.01160.1341-0.04080.143712.025-16.25422.503
35.761-3.06190.58184.6287-1.47912.6819-0.09950.0127-0.47230.07610.06810.6655-0.0594-0.22230.0190.15020.036-0.01340.1523-0.06180.2443-3.765-17.02718.67
41.16230.13430.22211.2978-0.8326.06980.00470.3101-0.3559-0.04350.1146-0.10530.33580.1797-0.10230.22550.04670.0130.2564-0.1130.250212.423-51.115-14.632
51.424-0.6132-0.57812.97710.24073.96480.05470.3611-0.08480.0294-0.0101-0.2436-0.16610.5134-0.00320.11610.00860.00560.206-0.03160.140112.684-43.437-9.204
68.17270.6954-3.42334.28641.13186.85920.4297-0.24550.4539-0.3963-0.2151-0.5206-0.38310.3939-0.15420.407-0.19160.14680.5607-0.05480.291718.438-34.085-21.181
75.0985-0.5311.16415.31480.5813.5086-0.0597-0.0307-0.41550.12810.22980.17720.07030.0142-0.1520.14310.00120.02620.1711-0.0120.13554.908-43.511-11.156
81.2621.1693-1.15491.687-0.43511.725-0.10180.676-0.0302-0.09480.33870.0414-0.31830.394-0.15710.2669-0.02390.07760.3254-0.04270.187311.064-41.995-18.439
91.04930.27031.71560.8361-0.57366.3861-0.06240.4744-0.1782-0.1720.1617-0.119-0.36620.2465-0.07180.23070.03820.03710.3509-0.09120.198516.792-44.929-13.759
102.57221.54311.19733.894-0.09552.7187-0.13540.16560.141-0.0567-0.0312-0.3583-0.39570.59520.14030.18-0.0424-0.010.28590.02130.205426.441-26.63415.228
111.43110.30560.73311.58640.51941.4896-0.0383-0.17620.03850.1345-0.0156-0.0886-0.00270.0420.0640.12080.0199-0.00670.1350.00850.109415.422-30.44717.802
121.90450.7267-0.45313.8953-0.04710.90860.0419-0.3928-0.57240.17360.0818-0.12840.2101-0.0189-0.09520.21970.0287-0.04370.23740.11310.260413.023-49.88519.39
131.1384-0.0227-0.19762.12460.16461.0365-0.0355-0.031-0.1846-0.09720.00560.0560.11180.0076-0.00540.10760.0140.00270.09760.00830.12711.385-38.67611.035
145.9215-4.23081.79388.1117-3.74053.8947-0.0642-0.0097-0.19920.0846-0.0380.5251-0.0778-0.18310.09040.15070.0650.02890.2497-0.04330.3068-10.047-11.29522.688
153.4381-1.27523.67693.6033-1.97874.4776-0.182-0.8242-0.15510.6320.26190.44520.33770.044-0.02920.41410.18360.12250.42620.05270.26-2.039-17.48831.801
164.2889-0.08222.00071.6966-2.22194.95150.35830.1948-0.1585-0.1127-0.01720.45720.18360.0654-0.28450.21420.0829-0.03370.1985-0.03650.21330.194-14.81520.59
174.20912.38874.32572.35311.59685.19620.08750.9191-0.2378-0.2727-0.0250.4012-0.06840.2548-0.04310.24780.168-0.07670.42-0.11980.30490.974-13.9115.477
182.79921.0326-3.13823.5511-0.36467.9032-0.0651-0.6480.30730.8309-0.0481-0.4481-0.041.03580.01480.28850.06-0.07670.308-0.11680.2519.485-11.07229.629
194.0961-0.1761-1.01363.6619-0.36595.1384-0.0978-0.61510.44310.5290.04340.1083-0.21320.00130.03960.24030.0786-0.0010.2322-0.07210.2409-0.901-8.01427.717
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1( CHAIN B AND RESID 84:99 )B84 - 99
2X-RAY DIFFRACTION2( CHAIN B AND RESID 100:110 )B100 - 110
3X-RAY DIFFRACTION3( CHAIN B AND RESID 111:127 )B111 - 127
4X-RAY DIFFRACTION4( CHAIN C AND RESID 1:30 )C1 - 30
5X-RAY DIFFRACTION5( CHAIN C AND RESID 31:39 )C31 - 39
6X-RAY DIFFRACTION6( CHAIN C AND RESID 40:45 )C40 - 45
7X-RAY DIFFRACTION7( CHAIN C AND RESID 46:81 )C46 - 81
8X-RAY DIFFRACTION8( CHAIN C AND RESID 82:98 )C82 - 98
9X-RAY DIFFRACTION9( CHAIN C AND RESID 99:116 )C99 - 116
10X-RAY DIFFRACTION10( CHAIN A AND RESID 334:370 )A334 - 370
11X-RAY DIFFRACTION11( CHAIN A AND RESID 371:459 )A371 - 459
12X-RAY DIFFRACTION12( CHAIN A AND RESID 460:479 )A460 - 479
13X-RAY DIFFRACTION13( CHAIN A AND RESID 480:517 )A480 - 517
14X-RAY DIFFRACTION14( CHAIN B AND RESID 1:17 )B1 - 17
15X-RAY DIFFRACTION15( CHAIN B AND RESID 18:32 )B18 - 32
16X-RAY DIFFRACTION16( CHAIN B AND RESID 33:39 )B33 - 39
17X-RAY DIFFRACTION17( CHAIN B AND RESID 40:51 )B40 - 51
18X-RAY DIFFRACTION18( CHAIN B AND RESID 52:60 )B52 - 60
19X-RAY DIFFRACTION19( CHAIN B AND RESID 61:83 )B61 - 83

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