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- EMDB-26838: Klebsiella pneumoniae adenosine monophosphate nucleosidase -

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Basic information

Entry
Database: EMDB / ID: EMD-26838
TitleKlebsiella pneumoniae adenosine monophosphate nucleosidase
Map dataSharpened map for building
Sample
  • Complex: Adenosine monophosphate nucleosidase
    • Protein or peptide: AMP nucleosidase
Keywordsnucleosidase / AMP / salvage / HYDROLASE
Function / homology
Function and homology information


AMP nucleosidase / AMP nucleosidase activity / nucleoside metabolic process / AMP salvage / cytoplasm
Similarity search - Function
AMP nucleoside phosphorylase, N-terminal / AMP nucleoside phosphorylase, N-terminal domain superfamily / Bacterial AMP nucleoside phosphorylase N-terminus / AMP nucleosidase / Nucleoside phosphorylase domain / Phosphorylase superfamily / Nucleoside phosphorylase superfamily
Similarity search - Domain/homology
Biological speciesKlebsiella pneumoniae (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.05 Å
AuthorsRichardson BC / French JB
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM124898 United States
CitationJournal: PLoS One / Year: 2022
Title: Structure of Klebsiella pneumoniae adenosine monophosphate nucleosidase.
Authors: Brian C Richardson / Roger Shek / Wesley C Van Voorhis / Jarrod B French /
Abstract: Klebsiella pneumoniae is a bacterial pathogen that is increasingly responsible for hospital-acquired pneumonia and sepsis. Progressive development of antibiotic resistance has led to higher mortality ...Klebsiella pneumoniae is a bacterial pathogen that is increasingly responsible for hospital-acquired pneumonia and sepsis. Progressive development of antibiotic resistance has led to higher mortality rates and creates a need for novel treatments. Because of the essential role that nucleotides play in many bacterial processes, enzymes involved in purine and pyrimidine metabolism and transport are ideal targets for the development of novel antibiotics. Herein we describe the structure of K. pneumoniae adenosine monophosphate nucleosidase (KpAmn), a purine salvage enzyme unique to bacteria, as determined by cryoelectron microscopy. The data detail a well conserved fold with a hexameric overall structure and clear density for the putative active site residues. Comparison to the crystal structures of homologous prokaryotic proteins confirms the presence of many of the conserved structural features of this protein yet reveals differences in distal loops in the absence of crystal contacts. This first cryo-EM structure of an Amn enzyme provides a basis for future structure-guided drug development and extends the accuracy of structural characterization of this family of proteins beyond this clinically relevant organism.
History
DepositionMay 3, 2022-
Header (metadata) releaseSep 28, 2022-
Map releaseSep 28, 2022-
UpdateJun 12, 2024-
Current statusJun 12, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26838.png

Downloads & links

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Map

FileDownload / File: emd_26838.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened map for building
Voxel sizeX=Y=Z: 0.8891 Å
Density
Contour LevelBy AUTHOR: 0.604
Minimum - Maximum-2.370475 - 3.7208796
Average (Standard dev.)0.0049120523 (±0.10807348)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 266.73 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_26838_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_26838_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_26838_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Adenosine monophosphate nucleosidase

EntireName: Adenosine monophosphate nucleosidase
Components
  • Complex: Adenosine monophosphate nucleosidase
    • Protein or peptide: AMP nucleosidase

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Supramolecule #1: Adenosine monophosphate nucleosidase

SupramoleculeName: Adenosine monophosphate nucleosidase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Klebsiella pneumoniae (bacteria)

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Macromolecule #1: AMP nucleosidase

MacromoleculeName: AMP nucleosidase / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO / EC number: AMP nucleosidase
Source (natural)Organism: Klebsiella pneumoniae (bacteria)
Molecular weightTheoretical: 55.326336 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MAHHHHHHMN IKAASLTPEQ ALAELEARYE ASVTALRKAI GDYIDHNTLP DTEARAEGLF VYPQLSVSWD GADHKALKTR AWGRFTHAG CYTTTITNPK LFRNYLLEQL TLLYQDYGAH ISVELSQHEI PYPYVIDGST LTLDRSMSAG LTRYFPTTEL S QIGDETAD ...String:
MAHHHHHHMN IKAASLTPEQ ALAELEARYE ASVTALRKAI GDYIDHNTLP DTEARAEGLF VYPQLSVSWD GADHKALKTR AWGRFTHAG CYTTTITNPK LFRNYLLEQL TLLYQDYGAH ISVELSQHEI PYPYVIDGST LTLDRSMSAG LTRYFPTTEL S QIGDETAD GLFHPTEFYP LSHFDARRVD FSLARLRHYT GTPAEHFQPY VLFTNYTRYV DEFVSWGCSQ ILDPDSPYIA LS CAGGIWI TAETEAPEQA ISDLAWKKHQ MPAWHLITHD GKGITLINIG VGPANAKTIC DHLAVLRPDV WLMIGHCGGL RES QAIGDY VLAHAYLRDD HVLDAVLPPD IPIPSIAEVQ RALYDATKQV SGMPGEEVKQ RLRTGTVVTT DDRNWELRYS ASAL RFNLS RAVAIDMESA TIAAQGYRFR VPYGTLLCVS DKPLHGEIKL PGQANRFYEG AISEHLQIGI RAIDLLRAEG DHMHS RKLR TFNEPPFR

UniProtKB: AMP nucleosidase

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration2 mg/mL
BufferpH: 7
Component:
ConcentrationNameFormula
25.0 uMHEPES
500.0 mMsodium chlorideNaClSodium chloride
5.0 %glycerol
2.0 mMDTT
0.025 %sodium azide
GridModel: C-flat-1.2/1.3 / Material: GOLD / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 96 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV / Details: 9 second blot.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.6 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 96000
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 679 / Average electron dose: 60.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1494578
Startup modelType of model: NONE
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.1)
Final 3D classificationNumber classes: 17 / Avg.num./class: 6752 / Software - Name: cryoSPARC (ver. 3.3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.1)
Final reconstructionNumber classes used: 17 / Applied symmetry - Point group: D3 (2x3 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 3.05 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.1) / Number images used: 114781
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

1t8r


Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-7uwq:
Klebsiella pneumoniae adenosine monophosphate nucleosidase

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