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- EMDB-36071: Membrane bound PRTase, C3 symmetry, donor bound -

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Basic information

Entry
Database: EMDB / ID: EMD-36071
TitleMembrane bound PRTase, C3 symmetry, donor bound
Map dataEM map of a membrane-bound PRTase, C3 symmetry donor-bound
Sample
  • Complex: trimeric phosphoribosyltransferase
    • Protein or peptide: Decaprenyl-phosphate phosphoribosyltransferase
  • Ligand: 1-O-pyrophosphono-5-O-phosphono-alpha-D-ribofuranose
  • Ligand: MAGNESIUM ION
  • Ligand: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
Keywordsphosphoribose transferase complex / MEMBRANE PROTEIN
Function / homology
Function and homology information


decaprenyl-phosphate phosphoribosyltransferase / arabinosyltransferase activity / glycolipid biosynthetic process / capsule polysaccharide biosynthetic process / transferase activity, transferring alkyl or aryl (other than methyl) groups / cell wall organization / magnesium ion binding / plasma membrane
Similarity search - Function
Prenyltransferase / UbiA prenyltransferase family / UbiA prenyltransferase superfamily / UbiA prenyltransferase family
Similarity search - Domain/homology
Decaprenyl-phosphate phosphoribosyltransferase
Similarity search - Component
Biological speciesMycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.76 Å
AuthorsWu FY / Gao S / Zhang L / Rao ZH
Funding support China, 1 items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, China)SRPG22-003 China
CitationJournal: Nat Microbiol / Year: 2024
Title: Structural analysis of phosphoribosyltransferase-mediated cell wall precursor synthesis in Mycobacterium tuberculosis.
Authors: Shan Gao / Fangyu Wu / Sudagar S Gurcha / Sarah M Batt / Gurdyal S Besra / Zihe Rao / Lu Zhang /
Abstract: In Mycobacterium tuberculosis, Rv3806c is a membrane-bound phosphoribosyltransferase (PRTase) involved in cell wall precursor production. It catalyses pentosyl phosphate transfer from phosphoribosyl ...In Mycobacterium tuberculosis, Rv3806c is a membrane-bound phosphoribosyltransferase (PRTase) involved in cell wall precursor production. It catalyses pentosyl phosphate transfer from phosphoribosyl pyrophosphate to decaprenyl phosphate, to generate 5-phospho-β-ribosyl-1-phosphoryldecaprenol. Despite Rv3806c being an attractive drug target, structural and molecular mechanistic insight into this PRTase is lacking. Here we report cryogenic electron microscopy structures for Rv3806c in the donor- and acceptor-bound states. In a lipidic environment, Rv3806c is trimeric, creating a UbiA-like fold. Each protomer forms two helical bundles, which, alongside the bound lipids, are required for PRTase activity in vitro. Mutational and functional analyses reveal that decaprenyl phosphate and phosphoribosyl pyrophosphate bind the intramembrane and extramembrane cavities of Rv3806c, respectively, in a distinct manner to that of UbiA superfamily enzymes. Our data suggest a model for Rv3806c-catalysed phosphoribose transfer through an inverting mechanism. These findings provide a structural basis for cell wall precursor biosynthesis that could have potential for anti-tuberculosis drug development.
History
DepositionMay 1, 2023-
Header (metadata) releaseFeb 7, 2024-
Map releaseFeb 7, 2024-
UpdateApr 17, 2024-
Current statusApr 17, 2024Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_36071.png

Downloads & links

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Map

FileDownload / File: emd_36071.map.gz / Format: CCP4 / Size: 113.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationEM map of a membrane-bound PRTase, C3 symmetry donor-bound
Voxel sizeX=Y=Z: 0.96 Å
Density
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-4.3568683 - 6.471416
Average (Standard dev.)0.0018321224 (±0.10137031)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions310310310
Spacing310310310
CellA=B=C: 297.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: EM half map B of a membrane-bound PRTase, C3 symmetry donor-bound

Fileemd_36071_half_map_1.map
AnnotationEM half map B of a membrane-bound PRTase, C3 symmetry donor-bound
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: EM half map A of a membrane-bound PRTase, C3 symmetry donor-bound

Fileemd_36071_half_map_2.map
AnnotationEM half map A of a membrane-bound PRTase, C3 symmetry donor-bound
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : trimeric phosphoribosyltransferase

EntireName: trimeric phosphoribosyltransferase
Components
  • Complex: trimeric phosphoribosyltransferase
    • Protein or peptide: Decaprenyl-phosphate phosphoribosyltransferase
  • Ligand: 1-O-pyrophosphono-5-O-phosphono-alpha-D-ribofuranose
  • Ligand: MAGNESIUM ION
  • Ligand: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate

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Supramolecule #1: trimeric phosphoribosyltransferase

SupramoleculeName: trimeric phosphoribosyltransferase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: trimeric phosphoribosyltransferase, donor bound
Source (natural)Organism: Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria)
Molecular weightTheoretical: 97.8 KDa

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Macromolecule #1: Decaprenyl-phosphate phosphoribosyltransferase

MacromoleculeName: Decaprenyl-phosphate phosphoribosyltransferase / type: protein_or_peptide / ID: 1 / Number of copies: 3 / Enantiomer: LEVO / EC number: decaprenyl-phosphate phosphoribosyltransferase
Source (natural)Organism: Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria)
Molecular weightTheoretical: 32.682355 KDa
Recombinant expressionOrganism: Mycolicibacterium smegmatis MC2 155 (bacteria)
SequenceString: MSEDVVTQPP ANLVAGVVKA IRPRQWVKNV LVLAAPLAAL GGGVRYDYVE VLSKVSMAFV VFSLAASAVY LVNDVRDVEA DREHPTKRF RPIAAGVVPE WLAYTVAVVL GVTSLAGAWM LTPNLALVMV VYLAMQLAYC FGLKHQAVVE ICVVSSAYLI R AIAGGVAT ...String:
MSEDVVTQPP ANLVAGVVKA IRPRQWVKNV LVLAAPLAAL GGGVRYDYVE VLSKVSMAFV VFSLAASAVY LVNDVRDVEA DREHPTKRF RPIAAGVVPE WLAYTVAVVL GVTSLAGAWM LTPNLALVMV VYLAMQLAYC FGLKHQAVVE ICVVSSAYLI R AIAGGVAT KIPLSKWFLL IMAFGSLFMV AGKRYAELHL AERTGAAIRK SLESYTSTYL RFVWTLSATA VVLCYGLWAF ER DGYSGSW FAVSMIPFTI AILRYAVDVD GGLAGEPEDI ALRDRVLQLL ALAWIATVGA AVAFG

UniProtKB: Decaprenyl-phosphate phosphoribosyltransferase

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Macromolecule #2: 1-O-pyrophosphono-5-O-phosphono-alpha-D-ribofuranose

MacromoleculeName: 1-O-pyrophosphono-5-O-phosphono-alpha-D-ribofuranose / type: ligand / ID: 2 / Number of copies: 3 / Formula: PRP
Molecular weightTheoretical: 390.07 Da
Chemical component information

ChemComp-PRP:
1-O-pyrophosphono-5-O-phosphono-alpha-D-ribofuranose

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Macromolecule #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 3 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #4: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]o...

MacromoleculeName: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
type: ligand / ID: 4 / Number of copies: 3 / Formula: PGW
Molecular weightTheoretical: 749.007 Da

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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

Concentration2.0 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
150.0 mMNaClsodium chloride
20.0 mMHepesHydroxyethylpiperazine Ethane Sulfonic Acid
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 40 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 %
DetailsThis sample was mono disperse

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Number real images: 9908 / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 130000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 3597451
Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.76 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4) / Number images used: 410532
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4)
Final 3D classificationNumber classes: 5 / Software - Name: cryoSPARC (ver. 4)

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