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- EMDB-14576: CRYO-EM STRUCTURE OF SARS-COV-2 SPIKE : H11-H4 Q98R H100E nanobod... -
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Open data
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Basic information
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Title | CRYO-EM STRUCTURE OF SARS-COV-2 SPIKE : H11-H4 Q98R H100E nanobody complex in 2Up1Down conformation | |||||||||
![]() | Sharpened map, output from postprocessing. | |||||||||
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Function / homology | ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() ![]() ![]() ![]() ![]() ![]() | |||||||||
Method | ![]() ![]() | |||||||||
![]() | Weckener M / Naismith JH | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Correlation between the binding affinity and the conformational entropy of nanobody SARS-CoV-2 spike protein complexes. Authors: Halina Mikolajek / Miriam Weckener / Z Faidon Brotzakis / Jiandong Huo / Evmorfia V Dalietou / Audrey Le Bas / Pietro Sormanni / Peter J Harrison / Philip N Ward / Steven Truong / Lucile ...Authors: Halina Mikolajek / Miriam Weckener / Z Faidon Brotzakis / Jiandong Huo / Evmorfia V Dalietou / Audrey Le Bas / Pietro Sormanni / Peter J Harrison / Philip N Ward / Steven Truong / Lucile Moynie / Daniel K Clare / Maud Dumoux / Joshua Dormon / Chelsea Norman / Naveed Hussain / Vinod Vogirala / Raymond J Owens / Michele Vendruscolo / James H Naismith / ![]() ![]() Abstract: Camelid single-domain antibodies, also known as nanobodies, can be readily isolated from naïve libraries for specific targets but often bind too weakly to their targets to be immediately useful. ...Camelid single-domain antibodies, also known as nanobodies, can be readily isolated from naïve libraries for specific targets but often bind too weakly to their targets to be immediately useful. Laboratory-based genetic engineering methods to enhance their affinity, termed maturation, can deliver useful reagents for different areas of biology and potentially medicine. Using the receptor binding domain (RBD) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein and a naïve library, we generated closely related nanobodies with micromolar to nanomolar binding affinities. By analyzing the structure-activity relationship using X-ray crystallography, cryoelectron microscopy, and biophysical methods, we observed that higher conformational entropy losses in the formation of the spike protein-nanobody complex are associated with tighter binding. To investigate this, we generated structural ensembles of the different complexes from electron microscopy maps and correlated the conformational fluctuations with binding affinity. This insight guided the engineering of a nanobody with improved affinity for the spike protein. | |||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 10.7 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 29.2 KB 29.2 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 10.7 KB | Display | ![]() |
Images | ![]() | 121 KB | ||
Masks | ![]() | 103 MB | ![]() | |
Others | ![]() ![]() ![]() ![]() | 80.8 MB 5.2 MB 80.8 MB 81.1 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 7z9rMC ![]() 7z1aC ![]() 7z1bC ![]() 7z1cC ![]() 7z1dC ![]() 7z1eC ![]() 7z6vC ![]() 7z7xC ![]() 7z85C ![]() 7z86C ![]() 7z9qC M: atomic model generated by this map C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | Sharpened map, output from postprocessing. | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.072 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
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Projections & Slices |
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Density Histograms |
-Additional map: Map from final 3D refinement.
File | emd_14576_additional_1.map | ||||||||||||
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Annotation | Map from final 3D refinement. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: Sharpened map, output from LocScale.
File | emd_14576_additional_2.map | ||||||||||||
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Annotation | Sharpened map, output from LocScale. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map from final 3D refinement.
File | emd_14576_half_map_1.map | ||||||||||||
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Annotation | Half map from final 3D refinement. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half map from final 3D refinement.
File | emd_14576_half_map_2.map | ||||||||||||
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Annotation | Half map from final 3D refinement. | ||||||||||||
Projections & Slices |
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Density Histograms |
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Sample components
-Entire : Complex between Spike and nanobody H11-H4 Q98R H100E
Entire | Name: Complex between Spike and nanobody H11-H4 Q98R H100E |
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Components |
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-Supramolecule #1: Complex between Spike and nanobody H11-H4 Q98R H100E
Supramolecule | Name: Complex between Spike and nanobody H11-H4 Q98R H100E / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: #1-#2 Details: The Spike protein is found in the 2Up1Down conformation. |
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Molecular weight | Theoretical: 15 KDa |
-Supramolecule #2: Spike glycoprotein trimer with fibritin tag
Supramolecule | Name: Spike glycoprotein trimer with fibritin tag / type: complex / Chimera: Yes / ID: 2 / Parent: 1 / Macromolecule list: #1 |
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Source (natural) | Organism: ![]() ![]() ![]() |
Recombinant expression | Organism: ![]() ![]() |
-Supramolecule #3: Nanobody H11-H4 Q98R H100E
Supramolecule | Name: Nanobody H11-H4 Q98R H100E / type: complex / Chimera: Yes / ID: 3 / Parent: 1 / Macromolecule list: #2 |
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Source (natural) | Organism: ![]() ![]() ![]() |
Recombinant expression | Organism: ![]() ![]() ![]() |
-Macromolecule #1: Spike glycoprotein,Fibritin
Macromolecule | Name: Spike glycoprotein,Fibritin / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 139.877906 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MFVFLVLLPL VSSQCVNLTT RTQLPPAYTN SFTRGVYYPD KVFRSSVLHS TQDLFLPFFS NVTWFHAIHV SGTNGTKRFD NPVLPFNDG VYFASTEKSN IIRGWIFGTT LDSKTQSLLI VNNATNVVIK VCEFQFCNDP FLGVYYHKNN KSWMESEFRV Y SSANNCTF ...String: MFVFLVLLPL VSSQCVNLTT RTQLPPAYTN SFTRGVYYPD KVFRSSVLHS TQDLFLPFFS NVTWFHAIHV SGTNGTKRFD NPVLPFNDG VYFASTEKSN IIRGWIFGTT LDSKTQSLLI VNNATNVVIK VCEFQFCNDP FLGVYYHKNN KSWMESEFRV Y SSANNCTF EYVSQPFLMD LEGKQGNFKN LREFVFKNID GYFKIYSKHT PINLVRDLPQ GFSALEPLVD LPIGINITRF QT LLALHRS YLTPGDSSSG WTAGAAAYYV GYLQPRTFLL KYNENGTITD AVDCALDPLS ETKCTLKSFT VEKGIYQTSN FRV QPTESI VRFPNITNLC PFGEVFNATR FASVYAWNRK RISNCVADYS VLYNSASFST FKCYGVSPTK LNDLCFTNVY ADSF VIRGD EVRQIAPGQT GKIADYNYKL PDDFTGCVIA WNSNNLDSKV GGNYNYLYRL FRKSNLKPFE RDISTEIYQA GSTPC NGVE GFNCYFPLQS YGFQPTNGVG YQPYRVVVLS FELLHAPATV CGPKKSTNLV KNKCVNFNFN GLTGTGVLTE SNKKFL PFQ QFGRDIADTT DAVRDPQTLE ILDITPCSFG GVSVITPGTN TSNQVAVLYQ DVNCTEVPVA IHADQLTPTW RVYSTGS NV FQTRAGCLIG AEHVNNSYEC DIPIGAGICA SYQTQTNSPG SASSVASQSI IAYTMSLGAE NSVAYSNNSI AIPTNFTI S VTTEILPVSM TKTSVDCTMY ICGDSTECSN LLLQYGSFCT QLNRALTGIA VEQDKNTQEV FAQVKQIYKT PPIKDFGGF NFSQILPDPS KPSKRSFIED LLFNKVTLAD AGFIKQYGDC LGDIAARDLI CAQKFNGLTV LPPLLTDEMI AQYTSALLAG TITSGWTFG AGAALQIPFA MQMAYRFNGI GVTQNVLYEN QKLIANQFNS AIGKIQDSLS STASALGKLQ DVVNQNAQAL N TLVKQLSS NFGAISSVLN DILSRLDPPE AEVQIDRLIT GRLQSLQTYV TQQLIRAAEI RASANLAATK MSECVLGQSK RV DFCGKGY HLMSFPQSAP HGVVFLHVTY VPAQEKNFTT APAICHDGKA HFPREGVFVS NGTHWFVTQR NFYEPQIITT DNT FVSGNC DVVIGIVNNT VYDPLQPELD SFKEELDKYF KNHTSPDVDL GDISGINASV VNIQKEIDRL NEVAKNLNES LIDL QELGK YEQGSGYIPE APRDGQAYVR KDGEWVLLST FLSLLNDIFE AQKIEWHEKH HHHHH |
-Macromolecule #2: Nanobody H11-H4 Q98R H100E
Macromolecule | Name: Nanobody H11-H4 Q98R H100E / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() ![]() |
Molecular weight | Theoretical: 14.107723 KDa |
Recombinant expression | Organism: ![]() ![]() ![]() |
Sequence | String: QVQLVESGGG LMQAGGSLRL SCAVSGRTFS TAAMGWFRQA PGKEREFVAA IRWSGGSAYY ADSVKGRFTI SRDKAKNTVY LQMNSLKYE DTAVYYCART EYVSYLLSDY ATWPYDYWGQ GTQVTVSS |
-Macromolecule #4: 2-acetamido-2-deoxy-beta-D-glucopyranose
Macromolecule | Name: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 4 / Number of copies: 26 / Formula: NAG |
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Molecular weight | Theoretical: 221.208 Da |
Chemical component information | ![]() ChemComp-NAG: |
-Experimental details
-Structure determination
Method | ![]() |
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Aggregation state | particle |
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Sample preparation
Concentration | 1 mg/mL | |||||||||
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Buffer | pH: 8 Component:
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Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Details: Grids were glow discharged twice, 60s each. | |||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 289.15 K / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | TFS KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD![]() |
Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Details | EPU auto-function coma free correction |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Average exposure time: 3.85 sec. / Average electron dose: 50.0 e/Å2 / Details: The images were collected as 50 frame movies |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |