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- PDB-8hx0: Cryo-EM structure of MsDps2 from Mycobacterium smegmatis -

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Basic information

Entry
Database: PDB / ID: 8hx0
TitleCryo-EM structure of MsDps2 from Mycobacterium smegmatis
ComponentsPutative starvation-induced DNA protecting protein/Ferritin and Dps
KeywordsDNA BINDING PROTEIN / MsDps2 / cryo-EM
Function / homology
Function and homology information


oxidoreductase activity, acting on metal ions / ferric iron binding / DNA binding
Similarity search - Function
Dps protein family signature 1. / DNA-binding protein Dps, conserved site / DNA-binding protein Dps / Ferritin/DPS protein domain / Ferritin-like domain / Ferritin-like / Ferritin-like superfamily
Similarity search - Domain/homology
Starvation-inducible DNA-binding protein or fine tangled pili major subunit
Similarity search - Component
Biological speciesMycolicibacterium smegmatis MC2 155 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsGarg, P. / Dutta, S.
Funding support India, 2items
OrganizationGrant numberCountry
Department of Biotechnology (DBT, India) India
Department of Science & Technology (DST, India) India
CitationJournal: J Mol Biol / Year: 2024
Title: Cryo-EM Reveals the Mechanism of DNA Compaction by Mycobacterium smegmatis Dps2.
Authors: Priyanka Garg / Thejas Satheesh / Mahipal Ganji / Somnath Dutta /
Abstract: DNA binding protein from starved cells (Dps) is a miniature ferritin complex, which plays a vital role in protecting bacterial DNA during starvation to maintain the integrity of bacteria under ...DNA binding protein from starved cells (Dps) is a miniature ferritin complex, which plays a vital role in protecting bacterial DNA during starvation to maintain the integrity of bacteria under hostile conditions. Several approaches, including cryo-electron tomography, have been previously implemented by other research groups to decipher the structure of the Dps protein bound to DNA. However, none of the structures of the Dps-DNA complex was resolved to high resolution to identify the DNA binding residues. Like other bacteria, Mycobacterium smegmatis also expresses Dps2 (called MsDps2), which binds DNA to protect it under oxidative stress conditions. In this study, we implemented various biochemical and biophysical studies to characterize the DNA protein interactions of Dps2 protein from Mycobacterium smegmatis. We employed single-particle cryo-EM-based structural analysis of MsDps2-DNA complexes and identified that the region close to the N-terminus confers the DNA binding property. Based on cryo-EM data, we also pinpointed several arginine residues, proximal to the DNA binding region, responsible for DNA binding. We also performed mutations of these residues, which dramatically reduced the MsDps2-DNA interaction. In addition, we proposed a model that elucidates the mechanism of DNA compaction, which adapts a lattice-like structure. We performed single-molecule imaging of MsDps2-DNA interactions that corroborate well with our structural studies. Taken together, our results delineate the specific MsDps2 residues that play an important role in DNA binding and compaction, providing new insights into Mycobacterial DNA compaction mechanisms under stress conditions.
History
DepositionJan 3, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jul 10, 2024Provider: repository / Type: Initial release
Revision 1.1Oct 16, 2024Group: Data collection / Database references / Structure summary
Category: citation / citation_author ...citation / citation_author / em_admin / pdbx_entry_details
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _em_admin.last_update
Revision 1.2Nov 13, 2024Group: Data collection / Database references / Category: citation / em_admin
Item: _citation.journal_volume / _citation.title / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Putative starvation-induced DNA protecting protein/Ferritin and Dps
B: Putative starvation-induced DNA protecting protein/Ferritin and Dps
C: Putative starvation-induced DNA protecting protein/Ferritin and Dps
D: Putative starvation-induced DNA protecting protein/Ferritin and Dps
E: Putative starvation-induced DNA protecting protein/Ferritin and Dps
G: Putative starvation-induced DNA protecting protein/Ferritin and Dps
I: Putative starvation-induced DNA protecting protein/Ferritin and Dps
K: Putative starvation-induced DNA protecting protein/Ferritin and Dps
F: Putative starvation-induced DNA protecting protein/Ferritin and Dps
H: Putative starvation-induced DNA protecting protein/Ferritin and Dps
J: Putative starvation-induced DNA protecting protein/Ferritin and Dps
L: Putative starvation-induced DNA protecting protein/Ferritin and Dps


Theoretical massNumber of molelcules
Total (without water)214,25812
Polymers214,25812
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Putative starvation-induced DNA protecting protein/Ferritin and Dps / MsDps2 / Starvation-inducible DNA-binding protein or fine tangled pili major subunit


Mass: 17854.863 Da / Num. of mol.: 12
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mycolicibacterium smegmatis MC2 155 (bacteria)
Gene: MSMEG_3242, MSMEI_3159 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0QXB7
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Cryo-EM structure of MsDps2 from Mycobacterium Smegmatis
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Mycolicibacterium smegmatis MC2 155 (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2250 nm / Nominal defocus min: 750 nm
Image recordingElectron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 98467 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00315147
ELECTRON MICROSCOPYf_angle_d0.520592
ELECTRON MICROSCOPYf_dihedral_angle_d4.3182082
ELECTRON MICROSCOPYf_chiral_restr0.0362415
ELECTRON MICROSCOPYf_plane_restr0.0042754

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