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- PDB-6w98: Single-Particle Cryo-EM Structure of Arabinofuranosyltransferase ... -
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Open data
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Basic information
Entry | Database: PDB / ID: 6w98 | |||||||||||||||||||||
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Title | Single-Particle Cryo-EM Structure of Arabinofuranosyltransferase AftD from Mycobacteria | |||||||||||||||||||||
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![]() | MEMBRANE PROTEIN / Glycosyltransferase / lipomannan / lipoarabinomannan / arabinofuranose / nanodisc / single-particle cryo-electron microscopy / acyl carrier protein | |||||||||||||||||||||
Function / homology | ![]() lipid biosynthetic process / lipid A biosynthetic process / phosphopantetheine binding / acyl binding / acyl carrier activity / fatty acid biosynthetic process / transferase activity / membrane => GO:0016020 / response to xenobiotic stimulus / lipid binding ...lipid biosynthetic process / lipid A biosynthetic process / phosphopantetheine binding / acyl binding / acyl carrier activity / fatty acid biosynthetic process / transferase activity / membrane => GO:0016020 / response to xenobiotic stimulus / lipid binding / cytoplasm / cytosol Similarity search - Function | |||||||||||||||||||||
Biological species | ![]() ![]() ![]() | |||||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å | |||||||||||||||||||||
![]() | Tan, Y.Z. / Zhang, L. / Rodrigues, J. / Zheng, R.B. / Giacometti, S.I. / Rosario, A.L. / Kloss, B. / Dandey, V.P. / Wei, H. / Brunton, R. ...Tan, Y.Z. / Zhang, L. / Rodrigues, J. / Zheng, R.B. / Giacometti, S.I. / Rosario, A.L. / Kloss, B. / Dandey, V.P. / Wei, H. / Brunton, R. / Raczkowski, A.M. / Athayde, D. / Catalao, M.J. / Pimentel, M. / Clarke, O.B. / Lowary, T.L. / Archer, M. / Niederweis, M. / Potter, C.S. / Carragher, B. / Mancia, F. | |||||||||||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Cryo-EM Structures and Regulation of Arabinofuranosyltransferase AftD from Mycobacteria. Authors: Yong Zi Tan / Lei Zhang / José Rodrigues / Ruixiang Blake Zheng / Sabrina I Giacometti / Ana L Rosário / Brian Kloss / Venkata P Dandey / Hui Wei / Richard Brunton / Ashleigh M Raczkowski ...Authors: Yong Zi Tan / Lei Zhang / José Rodrigues / Ruixiang Blake Zheng / Sabrina I Giacometti / Ana L Rosário / Brian Kloss / Venkata P Dandey / Hui Wei / Richard Brunton / Ashleigh M Raczkowski / Diogo Athayde / Maria João Catalão / Madalena Pimentel / Oliver B Clarke / Todd L Lowary / Margarida Archer / Michael Niederweis / Clinton S Potter / Bridget Carragher / Filippo Mancia / ![]() ![]() ![]() ![]() Abstract: Mycobacterium tuberculosis causes tuberculosis, a disease that kills over 1 million people each year. Its cell envelope is a common antibiotic target and has a unique structure due, in part, to two ...Mycobacterium tuberculosis causes tuberculosis, a disease that kills over 1 million people each year. Its cell envelope is a common antibiotic target and has a unique structure due, in part, to two lipidated polysaccharides-arabinogalactan and lipoarabinomannan. Arabinofuranosyltransferase D (AftD) is an essential enzyme involved in assembling these glycolipids. We present the 2.9-Å resolution structure of M. abscessus AftD, determined by single-particle cryo-electron microscopy. AftD has a conserved GT-C glycosyltransferase fold and three carbohydrate-binding modules. Glycan array analysis shows that AftD binds complex arabinose glycans. Additionally, AftD is non-covalently complexed with an acyl carrier protein (ACP). 3.4- and 3.5-Å structures of a mutant with impaired ACP binding reveal a conformational change, suggesting that ACP may regulate AftD function. Mutagenesis experiments using a conditional knockout constructed in M. smegmatis confirm the essentiality of the putative active site and the ACP binding for AftD function. | |||||||||||||||||||||
History |
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Structure visualization
Movie |
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 241.2 KB | Display | ![]() |
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PDB format | ![]() | 192.8 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1 MB | Display | ![]() |
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Full document | ![]() | 1.1 MB | Display | |
Data in XML | ![]() | 51.4 KB | Display | |
Data in CIF | ![]() | 77.7 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 21580MC ![]() 6wbxC ![]() 6wbyC M: map data used to model this data C: citing same article ( |
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Similar structure data | |
EM raw data | ![]() Data size: 2.7 TB Data #1: Unaligned and compressed multi-frame movies [micrographs - multiframe] Data #2: Aligned and dose-weighted micrographs [micrographs - single frame] Data #3: Final Particle Stacks with Refined Euler Angles and Shifts (Overall and Focused Refined on CBM3) [picked particles - single frame - processed]) |
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Links
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Assembly
Deposited unit | ![]()
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Components
-Protein , 2 types, 2 molecules AB
#1: Protein | Mass: 149584.281 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: SAMEA2161603_00457 / Production host: ![]() ![]() |
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#2: Protein | Mass: 8645.460 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
-Non-polymers , 4 types, 5 molecules ![](data/chem/img/CA.gif)
![](data/chem/img/6OU.gif)
![](data/chem/img/PNS.gif)
![](data/chem/img/HOH.gif)
![](data/chem/img/6OU.gif)
![](data/chem/img/PNS.gif)
![](data/chem/img/HOH.gif)
#3: Chemical | #4: Chemical | ChemComp-6OU / [( | #5: Chemical | ChemComp-PNS / | #6: Water | ChemComp-HOH / | |
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-Details
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 |
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Sample preparation
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Molecular weight |
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Source (natural) |
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Source (recombinant) | Organism: ![]() ![]() | ||||||||||||||||||||||||
Buffer solution | pH: 7.5 / Details: Solution was filtered and degassed. | ||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Protein was incorporated into lipid nanodiscs. | ||||||||||||||||||||||||
Specimen support | Grid material: COPPER / Grid type: Homemade | ||||||||||||||||||||||||
Vitrification | Instrument: SPOTITON / Cryogen name: ETHANE / Chamber temperature: 298 K Details: The Spotiton V1.0 robot was operating at room temperature and moderate humidity. |
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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: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / Alignment procedure: ZEMLIN TABLEAU |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 13.05 sec. / Electron dose: 95.72 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Num. of grids imaged: 4 / Num. of real images: 7274 |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 15 eV |
Image scans | Sampling size: 5 µm / Width: 3838 / Height: 3710 / Movie frames/image: 90 / Used frames/image: 1-90 |
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Processing
EM software |
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Image processing | Details: Energy filter slit width of 15 eV was used during the collection and was aligned automatically every hour using Leginon. | ||||||||||||||||||||||||||||||||||||||||||||||||||||
CTF correction | Details: CTF was estimated using GCTF and refined throughout the pipeline using cisTEM. Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 2800102 | ||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 490616 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Protocol: AB INITIO MODEL / Space: REAL |