+Open data
-Basic information
Entry | Database: PDB / ID: 8if8 | ||||||
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Title | Arabinosyltransferase AftA | ||||||
Components | Galactan 5-O-arabinofuranosyltransferase | ||||||
Keywords | MEMBRANE PROTEIN / Arabinosyltransferase / AftA / Mycobacterium tuberculosis | ||||||
Function / homology | Function and homology information galactan 5-O-arabinofuranosyltransferase / cell wall macromolecule biosynthetic process / UDP-galactosyltransferase activity / mycolate cell wall layer assembly / capsule polysaccharide biosynthetic process / glycosyltransferase activity / peptidoglycan-based cell wall / cell wall organization / plasma membrane Similarity search - Function | ||||||
Biological species | Mycobacterium tuberculosis (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å | ||||||
Authors | Gong, Y.C. / Rao, Z.H. / Zhang, L. | ||||||
Funding support | China, 1items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2023 Title: Structure of the priming arabinosyltransferase AftA required for AG biosynthesis of . Authors: Yicheng Gong / Chuancun Wei / Jun Wang / Nengjiang Mu / Qinhong Lu / Chengyao Wu / Ning Yan / Huifang Yang / Yao Zhao / Xiuna Yang / Sudagar S Gurcha / Natacha Veerapen / Sarah M Batt / ...Authors: Yicheng Gong / Chuancun Wei / Jun Wang / Nengjiang Mu / Qinhong Lu / Chengyao Wu / Ning Yan / Huifang Yang / Yao Zhao / Xiuna Yang / Sudagar S Gurcha / Natacha Veerapen / Sarah M Batt / Zhiqiang Hao / Lintai Da / Gurdyal S Besra / Zihe Rao / Lu Zhang / Abstract: Arabinogalactan (AG) is an essential cell wall component in mycobacterial species, including the deadly human pathogen . It plays a pivotal role in forming the rigid mycolyl-AG-peptidoglycan core for ...Arabinogalactan (AG) is an essential cell wall component in mycobacterial species, including the deadly human pathogen . It plays a pivotal role in forming the rigid mycolyl-AG-peptidoglycan core for in vitro growth. AftA is a membrane-bound arabinosyltransferase and a key enzyme involved in AG biosynthesis which bridges the assembly of the arabinan chain to the galactan chain. It is known that AftA catalyzes the transfer of the first arabinofuranosyl residue from the donor decaprenyl-monophosphoryl-arabinose to the mature galactan chain (i.e., priming); however, the priming mechanism remains elusive. Herein, we report the cryo-EM structure of AftA. The detergent-embedded AftA assembles as a dimer with an interface maintained by both the transmembrane domain (TMD) and the soluble C-terminal domain (CTD) in the periplasm. The structure shows a conserved glycosyltransferase-C fold and two cavities converging at the active site. A metal ion participates in the interaction of TMD and CTD of each AftA molecule. Structural analyses combined with functional mutagenesis suggests a priming mechanism catalyzed by AftA in AG biosynthesis. Our data further provide a unique perspective into anti-TB drug discovery. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8if8.cif.gz | 209.6 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8if8.ent.gz | 168.2 KB | Display | PDB format |
PDBx/mmJSON format | 8if8.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8if8_validation.pdf.gz | 367.9 KB | Display | wwPDB validaton report |
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Full document | 8if8_full_validation.pdf.gz | 377.2 KB | Display | |
Data in XML | 8if8_validation.xml.gz | 23.1 KB | Display | |
Data in CIF | 8if8_validation.cif.gz | 36.3 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/if/8if8 ftp://data.pdbj.org/pub/pdb/validation_reports/if/8if8 | HTTPS FTP |
-Related structure data
Related structure data | 35410MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 69576.938 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Mycobacterium tuberculosis (strain ATCC 25618 / H37Rv) (bacteria) Gene: aftA, Rv3792 / Plasmid: PMV261 Production host: Mycolicibacterium smegmatis MC2 155 (bacteria) Strain (production host): MC2 155 References: UniProt: P9WN03, galactan 5-O-arabinofuranosyltransferase #2: 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 | Name: AftA dimer / Type: COMPLEX / Details: AftA subunit1, AftA subunit2 / Entity ID: #1 / Source: RECOMBINANT |
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Molecular weight | Value: 0.0695 MDa / Experimental value: NO |
Source (natural) | Organism: Mycobacterium tuberculosis H37Rv (bacteria) |
Source (recombinant) | Organism: Mycolicibacterium smegmatis MC2 155 (bacteria) / Strain: MC2 155 |
Buffer solution | pH: 8 / Details: 150mM NaCl, 20mM Hepes, 0.04%GDN |
Buffer component | Conc.: 150 mM / Name: sodium chloride / Formula: NaCl |
Specimen | Conc.: 6.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: the sample was mono disperse |
Specimen support | Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 281 K |
-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: SPOT SCAN |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 10500 X / Calibrated magnification: 59000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: BASIC |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 2.4 sec. / Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 6113 / Num. of real images: 6113 / Details: images were collected in high resolution mode |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
-Processing
Software | Name: PHENIX / Version: 1.18.2_3874: / Classification: refinement | ||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Particle selection | Num. of particles selected: 2752481 | ||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 147896 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||
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