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- PDB-8x8o: Cryo-EM structure of a bacteriophage tail- spike protein against ... -

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Basic information

Entry
Database: PDB / ID: 8x8o
TitleCryo-EM structure of a bacteriophage tail- spike protein against Klebsiella pneumoniae K64,ORF41(K64-ORF41) in 5 mM EDTA
ComponentsProbable tail spike protein
KeywordsLYASE / K64-ORF41 / depolymerase / cryo-EM / Klebsiella pneumoniae K64 / capsule polysaccharide / EDTA
Function / homology
Function and homology information


symbiont entry into host cell via disruption of host cell glycocalyx / symbiont entry into host cell via disruption of host cell envelope / virus tail / outer membrane / adhesion receptor-mediated virion attachment to host cell
Similarity search - Function
Tail spike TSP1/Gp66, N-terminal domain / Tail spike TSP1/Gp66 receptor binding N-terminal domain / Bacteriophage T7 tail fibre protein / Phage T7 tail fibre protein / Trimeric autotransporter adhesin YadA-like, stalk domain / Coiled stalk of trimeric autotransporter adhesin / Serralysin-like metalloprotease, C-terminal
Similarity search - Domain/homology
Probable tail spike protein
Similarity search - Component
Biological speciesKlebsiella phage SH-Kp 152410 (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.5 Å
AuthorsXie, Y. / Huang, T. / Zhang, Z. / Tao, X.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32200803 China
CitationJournal: Int J Biol Macromol / Year: 2024
Title: Structural and functional basis of bacteriophage K64-ORF41 depolymerase for capsular polysaccharide degradation of Klebsiella pneumoniae K64.
Authors: Tianyun Huang / Zhuoyuan Zhang / Xin Tao / Xinyu Shi / Peng Lin / Dan Liao / Chenyu Ma / Xinle Cai / Wei Lin / Xiaofan Jiang / Peng Luo / Shan Wu / Yuan Xie /
Abstract: Capsule polysaccharide is an important virulence factor of Klebsiella pneumoniae (K. pneumoniae), which protects bacteria against the host immune response. A promising therapeutic approach is using ...Capsule polysaccharide is an important virulence factor of Klebsiella pneumoniae (K. pneumoniae), which protects bacteria against the host immune response. A promising therapeutic approach is using phage-derived depolymerases to degrade the capsular polysaccharide and expose and sensitize the bacteria to the host immune system. Here we determined the cryo-electron microscopy (cryo-EM) structures of a bacteriophage tail-spike protein against K. pneumoniae K64, ORF41 (K64-ORF41) and ORF41 in EDTA condition (K64-ORF41), at 2.37 Å and 2.50 Å resolution, respectively, for the first time. K64-ORF41 exists as a trimer and each protomer contains a β-helix domain including a right-handed parallel β-sheet helix fold capped at both ends, an insertion domain, and one β-sheet jellyroll domain. Moreover, our structural comparison with other depolymerases of K. pneumoniae suggests that the catalytic residues (Tyr528, His574 and Arg628) are highly conserved although the substrate of capsule polysaccharide is variable. Besides that, we figured out the important residues involved in the substrate binding pocket including Arg405, Tyr526, Trp550 and Phe669. This study establishes the structural and functional basis for the promising phage-derived broad-spectrum activity depolymerase therapeutics and effective CPS-degrading agents for the treatment of carbapenem-resistant K. pneumoniae K64 infections.
History
DepositionNov 27, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Sep 4, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Probable tail spike protein
B: Probable tail spike protein
C: Probable tail spike protein


Theoretical massNumber of molelcules
Total (without water)336,7553
Polymers336,7553
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Probable tail spike protein


Mass: 112251.500 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Klebsiella phage SH-Kp 152410 (virus) / Gene: 2410_orf00041 / Production host: Escherichia coli (E. coli) / References: UniProt: A0A2K9VGS2

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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: K64-ORF41 in 5 mM EDTA / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Klebsiella phage SH-Kp 152410 (virus)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

MicroscopyModel: FEI TITAN
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1500 nm / Nominal defocus min: 1100 nm
Image recordingElectron dose: 54 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

CTF correctionType: NONE
3D reconstructionResolution: 2.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 371259 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00318090
ELECTRON MICROSCOPYf_angle_d0.50424636
ELECTRON MICROSCOPYf_dihedral_angle_d4.9492529
ELECTRON MICROSCOPYf_chiral_restr0.0462742
ELECTRON MICROSCOPYf_plane_restr0.0043225

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