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Yorodumi- EMDB-27903: Design of Diverse Asymmetric Pockets in de novo Homo-oligomeric P... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-27903 | |||||||||
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Title | Design of Diverse Asymmetric Pockets in de novo Homo-oligomeric Proteins | |||||||||
Map data | Sharpened Map | |||||||||
Sample |
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Keywords | tetramer / oligomer / pockets / de novo design / rosetta / cryoEM / DE NOVO PROTEIN | |||||||||
Biological species | Escherichia coli (E. coli) / synthetic construct (others) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.85 Å | |||||||||
Authors | Gerben S / Borst AJ / Baker D | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Biochemistry / Year: 2023 Title: Design of Diverse Asymmetric Pockets in Homo-oligomeric Proteins. Authors: Stacey R Gerben / Andrew J Borst / Derrick R Hicks / Isabelle Moczygemba / David Feldman / Brian Coventry / Wei Yang / Asim K Bera / Marcos Miranda / Alex Kang / Hannah Nguyen / David Baker / Abstract: A challenge for design of protein-small-molecule recognition is that incorporation of cavities with size, shape, and composition suitable for specific recognition can considerably destabilize protein ...A challenge for design of protein-small-molecule recognition is that incorporation of cavities with size, shape, and composition suitable for specific recognition can considerably destabilize protein monomers. This challenge can be overcome through binding pockets formed at homo-oligomeric interfaces between folded monomers. Interfaces surrounding the central homo-oligomer symmetry axes necessarily have the same symmetry and so may not be well suited to binding asymmetric molecules. To enable general recognition of arbitrary asymmetric substrates and small molecules, we developed an approach to designing asymmetric interfaces at off-axis sites on homo-oligomers, analogous to those found in native homo-oligomeric proteins such as glutamine synthetase. We symmetrically dock curved helical repeat proteins such that they form pockets at the asymmetric interface of the oligomer with sizes ranging from several angstroms, appropriate for binding a single ion, to up to more than 20 Å across. Of the 133 proteins tested, 84 had soluble expression in , 47 had correct oligomeric states in solution, 35 had small-angle X-ray scattering (SAXS) data largely consistent with design models, and 8 had negative-stain electron microscopy (nsEM) 2D class averages showing the structures coming together as designed. Both an X-ray crystal structure and a cryogenic electron microscopy (cryoEM) structure are close to the computational design models. The nature of these proteins as homo-oligomers allows them to be readily built into higher-order structures such as nanocages, and the asymmetric pockets of these structures open rich possibilities for small-molecule binder design free from the constraints associated with monomer destabilization. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_27903.map.gz | 38.1 MB | EMDB map data format | |
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Header (meta data) | emd-27903-v30.xml emd-27903.xml | 16.9 KB 16.9 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_27903_fsc.xml | 7.3 KB | Display | FSC data file |
Images | emd_27903.png | 82.4 KB | ||
Filedesc metadata | emd-27903.cif.gz | 5.2 KB | ||
Others | emd_27903_additional_1.map.gz emd_27903_half_map_1.map.gz emd_27903_half_map_2.map.gz | 19.4 MB 37.5 MB 37.6 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-27903 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-27903 | HTTPS FTP |
-Validation report
Summary document | emd_27903_validation.pdf.gz | 750.5 KB | Display | EMDB validaton report |
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Full document | emd_27903_full_validation.pdf.gz | 750.1 KB | Display | |
Data in XML | emd_27903_validation.xml.gz | 15 KB | Display | |
Data in CIF | emd_27903_validation.cif.gz | 19.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-27903 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-27903 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_27903.map.gz / Format: CCP4 / Size: 40.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Sharpened Map | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.84 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Additional map: Unsharpened Map
File | emd_27903_additional_1.map | ||||||||||||
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Annotation | Unsharpened Map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half Map A
File | emd_27903_half_map_1.map | ||||||||||||
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Annotation | Half Map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Half Map B
File | emd_27903_half_map_2.map | ||||||||||||
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Annotation | Half Map B | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : SG135
Entire | Name: SG135 |
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Components |
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-Supramolecule #1: SG135
Supramolecule | Name: SG135 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Escherichia coli (E. coli) |
-Macromolecule #1: SG135
Macromolecule | Name: SG135 / type: protein_or_peptide / ID: 1 Details: Deleted loop consisting of residues 173-179 due to lack of confident map density Number of copies: 4 / Enantiomer: LEVO |
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Source (natural) | Organism: synthetic construct (others) |
Molecular weight | Theoretical: 23.521809 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | String: DREIKEEARK LIREAIELLQ KGDPRAKEIL RQAILILLAI RLLEEMEENI EKAEKLGNEE LSELAKRAIK LVREALELLK EGDPRAEEI LKLALKIIKA ILLLLEMYEN IKQAEELGDE DLSELAKIAI RLVRQALKLL QEGDPRAEEI LEIALRIIKL I LQLLFLKQ ...String: DREIKEEARK LIREAIELLQ KGDPRAKEIL RQAILILLAI RLLEEMEENI EKAEKLGNEE LSELAKRAIK LVREALELLK EGDPRAEEI LKLALKIIKA ILLLLEMYEN IKQAEELGDE DLSELAKIAI RLVRQALKLL QEGDPRAEEI LEIALRIIKL I LQLLFLKQ RIEEAKKKGD QQFVFEAEEK IRRIVEELFK LLEG |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1.0 mg/mL |
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Buffer | pH: 7.5 |
Grid | Model: C-flat-1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 63.56 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.7 µm / Nominal defocus min: 0.8 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |