+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-16917 | ||||||||||||
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Title | Human apo pseudouridine synthase 3 (PUS3) | ||||||||||||
Map data | human full-legnth apo PUS3 | ||||||||||||
Sample |
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Keywords | RNA modification / pseudouridylation / tRNA / homodimer / RNA BINDING PROTEIN | ||||||||||||
Function / homology | Function and homology information tRNA pseudouridine38/39 synthase / tRNA pseudouridine synthase activity / tRNA pseudouridine synthesis / mRNA pseudouridine synthesis / pseudouridine synthase activity / tRNA modification in the nucleus and cytosol / tRNA modification / RNA binding / nucleus / cytoplasm / cytosol Similarity search - Function | ||||||||||||
Biological species | Homo sapiens (human) | ||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 6.5 Å | ||||||||||||
Authors | Lin T-Y / Glatt S / Koziej L | ||||||||||||
Funding support | Poland, European Union, Switzerland, 3 items
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Citation | Journal: Mol Cell / Year: 2024 Title: The molecular basis of tRNA selectivity by human pseudouridine synthase 3. Authors: Ting-Yu Lin / Leon Kleemann / Jakub Jeżowski / Dominika Dobosz / Michał Rawski / Paulina Indyka / Grzegorz Ważny / Rahul Mehta / Andrzej Chramiec-Głąbik / Łukasz Koziej / Tristan Ranff ...Authors: Ting-Yu Lin / Leon Kleemann / Jakub Jeżowski / Dominika Dobosz / Michał Rawski / Paulina Indyka / Grzegorz Ważny / Rahul Mehta / Andrzej Chramiec-Głąbik / Łukasz Koziej / Tristan Ranff / Christian Fufezan / Mateusz Wawro / Jakub Kochan / Joanna Bereta / Sebastian A Leidel / Sebastian Glatt / Abstract: Pseudouridine (Ψ), the isomer of uridine, is ubiquitously found in RNA, including tRNA, rRNA, and mRNA. Human pseudouridine synthase 3 (PUS3) catalyzes pseudouridylation of position 38/39 in tRNAs. ...Pseudouridine (Ψ), the isomer of uridine, is ubiquitously found in RNA, including tRNA, rRNA, and mRNA. Human pseudouridine synthase 3 (PUS3) catalyzes pseudouridylation of position 38/39 in tRNAs. However, the molecular mechanisms by which it recognizes its RNA targets and achieves site specificity remain elusive. Here, we determine single-particle cryo-EM structures of PUS3 in its apo form and bound to three tRNAs, showing how the symmetric PUS3 homodimer recognizes tRNAs and positions the target uridine next to its active site. Structure-guided and patient-derived mutations validate our structural findings in complementary biochemical assays. Furthermore, we deleted PUS1 and PUS3 in HEK293 cells and mapped transcriptome-wide Ψ sites by Pseudo-seq. Although PUS1-dependent sites were detectable in tRNA and mRNA, we found no evidence that human PUS3 modifies mRNAs. Our work provides the molecular basis for PUS3-mediated tRNA modification in humans and explains how its tRNA modification activity is linked to intellectual disabilities. | ||||||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_16917.map.gz | 30.8 MB | EMDB map data format | |
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Header (meta data) | emd-16917-v30.xml emd-16917.xml | 17.5 KB 17.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_16917_fsc.xml | 8.5 KB | Display | FSC data file |
Images | emd_16917.png | 26.8 KB | ||
Masks | emd_16917_msk_1.map | 64 MB | Mask map | |
Filedesc metadata | emd-16917.cif.gz | 5.4 KB | ||
Others | emd_16917_half_map_1.map.gz emd_16917_half_map_2.map.gz | 59.2 MB 59.2 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-16917 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-16917 | HTTPS FTP |
-Validation report
Summary document | emd_16917_validation.pdf.gz | 672.7 KB | Display | EMDB validaton report |
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Full document | emd_16917_full_validation.pdf.gz | 672.3 KB | Display | |
Data in XML | emd_16917_validation.xml.gz | 16.3 KB | Display | |
Data in CIF | emd_16917_validation.cif.gz | 21 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-16917 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-16917 | HTTPS FTP |
-Related structure data
Related structure data | 9f9qMC 8okdC 9enbC 9encC 9eneC 9enfC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_16917.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | human full-legnth apo PUS3 | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.86 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_16917_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: human full-legnth apo PUS3 half map A
File | emd_16917_half_map_1.map | ||||||||||||
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Annotation | human full-legnth apo PUS3 half map A | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: human full-legnth apo PUS3 half map B
File | emd_16917_half_map_2.map | ||||||||||||
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Annotation | human full-legnth apo PUS3 half map B | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : homodimer of PUS3
Entire | Name: homodimer of PUS3 |
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Components |
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-Supramolecule #1: homodimer of PUS3
Supramolecule | Name: homodimer of PUS3 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 550 KDa |
-Macromolecule #1: pseudouridine syntase 3
Macromolecule | Name: pseudouridine syntase 3 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO / EC number: tRNA pseudouridine38/39 synthase |
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Source (natural) | Organism: Homo sapiens (human) |
Recombinant expression | Organism: Spodoptera frugiperda (fall armyworm) |
Sequence | String: AMADNDTDRN QTEKLLKRVR ELEQEVQRLK KEQAKNKEDS NIRENSAGAG KTKRAFDFSA HGRRHVALRI AYMGWGYQGF ASQENTNNTI EEKLFEALTK TRLVESRQTS NYHRCGRTAK GVSAFGQVIS LDLRSQFPRG RDSEDFNVKE EANAAAEEIR YTHILNRVLP ...String: AMADNDTDRN QTEKLLKRVR ELEQEVQRLK KEQAKNKEDS NIRENSAGAG KTKRAFDFSA HGRRHVALRI AYMGWGYQGF ASQENTNNTI EEKLFEALTK TRLVESRQTS NYHRCGRTAK GVSAFGQVIS LDLRSQFPRG RDSEDFNVKE EANAAAEEIR YTHILNRVLP PDIRILAWAP VEPSFSARFS CLERTYRYFF PRADLDIVTM DYAAQKYVGT HDFRNLCKMD VANGVINFQR TILSAQVQLV GQSPGEGRWQ EPFQLCQFEV TGQAFLYHQV RCMMAILFLI GQGMEKPEII DELLNIEKNP QKPQYSMAVE FPLVLYDCKF ENVKWIYDQE AQEFNITHLQ QLWANHAVKT HMLYSMLQGL DTVPVPCGIG PKMDGMTEWG NVKPSVIKQT SAFVEGVKMR TYKPLMDRPK CQGLESRIQH FVRRGRIEHP HLFHEEETKA KRDCNDTLEE ENTNLETPTK RVCVDTEIKS II UniProtKB: tRNA pseudouridine(38/39) synthase |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.3 mg/mL |
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Buffer | pH: 7.5 |
Grid | Model: C-flat-2/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR / Details: 8 mA |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | TFS KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 7353 / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.8000000000000003 µm / Nominal defocus min: 3.0 µm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |