+Open data
-Basic information
Entry | Database: PDB / ID: 7b5y | |||||||||
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Title | S. agalactiae BusR in complex with its busAB-promotor DNA | |||||||||
Components |
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Keywords | DNA BINDING PROTEIN / Repressor / complex / GntR | |||||||||
Function / homology | Function and homology information monoatomic cation transmembrane transporter activity / potassium ion transport / DNA-binding transcription factor activity Similarity search - Function | |||||||||
Biological species | Streptococcus agalactiae (bacteria) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 7.1 Å | |||||||||
Authors | Bandera, A.M. / Witte, G. | |||||||||
Funding support | Germany, 2items
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Citation | Journal: Nucleic Acids Res / Year: 2021 Title: BusR senses bipartite DNA binding motifs by a unique molecular ruler architecture. Authors: Adrian M Bandera / Joseph Bartho / Katja Lammens / David Jan Drexler / Jasmin Kleinschwärzer / Karl-Peter Hopfner / Gregor Witte / Abstract: The cyclic dinucleotide second messenger c-di-AMP is a major player in regulation of potassium homeostasis and osmolyte transport in a variety of bacteria. Along with various direct interactions with ...The cyclic dinucleotide second messenger c-di-AMP is a major player in regulation of potassium homeostasis and osmolyte transport in a variety of bacteria. Along with various direct interactions with proteins such as potassium channels, the second messenger also specifically binds to transcription factors, thereby altering the processes in the cell on the transcriptional level. We here describe the structural and biochemical characterization of BusR from the human pathogen Streptococcus agalactiae. BusR is a member of a yet structurally uncharacterized subfamily of the GntR family of transcription factors that downregulates transcription of the genes for the BusA (OpuA) glycine-betaine transporter upon c-di-AMP binding. We report crystal structures of full-length BusR, its apo and c-di-AMP bound effector domain, as well as cryo-EM structures of BusR bound to its operator DNA. Our structural data, supported by biochemical and biophysical data, reveal that BusR utilizes a unique domain assembly with a tetrameric coiled-coil in between the binding platforms, serving as a molecular ruler to specifically recognize a 22 bp separated bipartite binding motif. Binding of c-di-AMP to BusR induces a shift in equilibrium from an inactivated towards an activated state that allows BusR to bind the target DNA, leading to transcriptional repression. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7b5y.cif.gz | 198.2 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7b5y.ent.gz | 153.3 KB | Display | PDB format |
PDBx/mmJSON format | 7b5y.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7b5y_validation.pdf.gz | 1 MB | Display | wwPDB validaton report |
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Full document | 7b5y_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 7b5y_validation.xml.gz | 40.2 KB | Display | |
Data in CIF | 7b5y_validation.cif.gz | 60.1 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/b5/7b5y ftp://data.pdbj.org/pub/pdb/validation_reports/b5/7b5y | HTTPS FTP |
-Related structure data
Related structure data | 12051MC 7b5tC 7b5uC 7b5wC 7oz3C C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | |
Other databases |
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-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 23880.160 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Details: N-terminal residues GP result from purification tag Source: (gene. exp.) Streptococcus agalactiae (bacteria) Gene: BM110_ORF1201, AX245_01365, C6N10_09995, F5043_05515, GD434_05225, RDF_1124 Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: K0JNC6 #2: DNA chain | | Mass: 14309.208 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Streptococcus agalactiae (bacteria) #3: DNA chain | | Mass: 14020.027 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Streptococcus agalactiae (bacteria) #4: Chemical | Has ligand of interest | Y | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component |
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Molecular weight | Value: 0.123 MDa / Experimental value: YES | ||||||||||||||||||||||||
Source (natural) |
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Source (recombinant) |
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Buffer solution | pH: 6.5 / Details: degassed, filtered | ||||||||||||||||||||||||
Buffer component | Conc.: 20 mM / Name: hepes / Formula: HEPES | ||||||||||||||||||||||||
Specimen | Conc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: homogeneous and monodisperse sample | ||||||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 200 divisions/in. / Grid type: UltrAuFoil R2/2 | ||||||||||||||||||||||||
Vitrification | Instrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 283 K Details: 0.05% beta-octyl glycoside added prior to plunge freezing |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD |
Image recording | Electron dose: 45 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 370421 / Details: CryoSparc blob-picker | ||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||
3D reconstruction | Resolution: 7.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 20342 / Symmetry type: POINT | ||||||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL | ||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 7B5T Accession code: 7B5T / Source name: PDB / Type: experimental model |