[English] 日本語
![](img/lk-miru.gif)
- EMDB-33677: CryoEM structure of type III-E CRISPR gRAMP-crRNA complex bound t... -
+
Open data
-
Basic information
Entry | ![]() | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | CryoEM structure of type III-E CRISPR gRAMP-crRNA complex bound to non-self RNA target | |||||||||
![]() | ||||||||||
![]() |
| |||||||||
![]() | Nuclease / STRUCTURAL PROTEIN-RNA COMPLEX | |||||||||
Function / homology | : / CRISPR type III-associated protein / RAMP superfamily / defense response to virus / RNA binding / RAMP superfamily protein![]() | |||||||||
Biological species | ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.54 Å | |||||||||
![]() | Zhang JT / Cui N / Huang HD / Jia N | |||||||||
Funding support | 1 items
| |||||||||
![]() | ![]() Title: Structural basis for the non-self RNA-activated protease activity of the type III-E CRISPR nuclease-protease Craspase. Authors: Ning Cui / Jun-Tao Zhang / Zhuolin Li / Xiao-Yu Liu / Chongyuan Wang / Hongda Huang / Ning Jia / ![]() Abstract: The RNA-targeting type III-E CRISPR-gRAMP effector interacts with a caspase-like protease TPR-CHAT to form the CRISPR-guided caspase complex (Craspase), but their functional mechanism is unknown. ...The RNA-targeting type III-E CRISPR-gRAMP effector interacts with a caspase-like protease TPR-CHAT to form the CRISPR-guided caspase complex (Craspase), but their functional mechanism is unknown. Here, we report cryo-EM structures of the type III-E gRAMP and gRAMP-TPR-CHAT complexes, before and after either self or non-self RNA target binding, and elucidate the mechanisms underlying RNA-targeting and non-self RNA-induced protease activation. The associated TPR-CHAT adopted a distinct conformation upon self versus non-self RNA target binding, with nucleotides at positions -1 and -2 of the CRISPR-derived RNA (crRNA) serving as a sensor. Only binding of the non-self RNA target activated the TPR-CHAT protease, leading to cleavage of Csx30 protein. Furthermore, TPR-CHAT structurally resembled eukaryotic separase, but with a distinct mechanism for protease regulation. Our findings should facilitate the development of gRAMP-based RNA manipulation tools, and advance our understanding of the virus-host discrimination process governed by a nuclease-protease Craspase during type III-E CRISPR-Cas immunity. | |||||||||
History |
|
-
Structure visualization
Supplemental images |
---|
-
Downloads & links
-EMDB archive
Map data | ![]() | 59.7 MB | ![]() | |
---|---|---|---|---|
Header (meta data) | ![]() ![]() | 18 KB 18 KB | Display Display | ![]() |
Images | ![]() | 75.3 KB | ||
Masks | ![]() | 64 MB | ![]() | |
Filedesc metadata | ![]() | 6.6 KB | ||
Others | ![]() ![]() | 59.4 MB 59.4 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 7y81MC ![]() 7y80C ![]() 7y82C ![]() 7y83C ![]() 7y84C ![]() 7y85C M: atomic model generated by this map C: citing same article ( |
---|---|
Similar structure data | Similarity search - Function & homology ![]() |
-
Links
EMDB pages | ![]() ![]() |
---|
-
Map
File | ![]() | ||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Voxel size | X=Y=Z: 1.1 Å | ||||||||||||||||||||
Density |
| ||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
|
-Supplemental data
-Mask #1
File | ![]() | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: #2
File | emd_33677_half_map_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: #1
File | emd_33677_half_map_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-
Sample components
-Entire : Type III-E CRISPR gRAMP-crRNA binary complex
Entire | Name: Type III-E CRISPR gRAMP-crRNA binary complex |
---|---|
Components |
|
-Supramolecule #1: Type III-E CRISPR gRAMP-crRNA binary complex
Supramolecule | Name: Type III-E CRISPR gRAMP-crRNA binary complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3 |
---|---|
Source (natural) | Organism: ![]() |
-Supramolecule #2: gRAMP
Supramolecule | Name: gRAMP / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1 |
---|---|
Source (natural) | Organism: ![]() |
-Supramolecule #3: crRNA
Supramolecule | Name: crRNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2 |
---|
-Supramolecule #4: Non-self RNA target
Supramolecule | Name: Non-self RNA target / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #3 |
---|
-Macromolecule #1: RAMP superfamily protein
Macromolecule | Name: RAMP superfamily protein / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO |
---|---|
Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 198.652688 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MHHHHHHKSN DMNITVELTF FEPYRLVEWF DWDARKKSHS AMRGQAFAQW TWKGKGRTAG KSFITGTLVR SAVIKAVEEL LSLNNGKWE GVPCCNGSFQ TDESKGKKPS FLRKRHTLQW QANNKNICDK EEACPFCILL GRFDNAGKVH ERNKDYDIHF S NFDLDHKQ ...String: MHHHHHHKSN DMNITVELTF FEPYRLVEWF DWDARKKSHS AMRGQAFAQW TWKGKGRTAG KSFITGTLVR SAVIKAVEEL LSLNNGKWE GVPCCNGSFQ TDESKGKKPS FLRKRHTLQW QANNKNICDK EEACPFCILL GRFDNAGKVH ERNKDYDIHF S NFDLDHKQ EKNDLRLVDI ASGRILNRVD FDTGKAKDYF RTWEADYETY GTYTGRITLR NEHAKKLLLA SLGFVDKLCG AL CRIEVIK KSESPLPSDT KEQSYTKDDT VEVLSEDHND ELRKQAEVIV EAFKQNDKLE KIRILADAIR TLRLHGEGVI EKD ELPDGK EERDKGHHLW DIKVQGTALR TKLKELWQSN KDIGWRKFTE MLGSNLYLIY KKETGGVSTR FRILGDTEYY SKAH DSEGS DLFIPVTPPE GIETKEWIIV GRLKAATPFY FGVQQPSDSI PGKEKKSEDS LVINEHTSFN ILLDKENRYR IPRSA LRGA LRRDLRTAFG SGCNVSLGGQ ILCNCKVCIE MRRITLKDSV SDFSEPPEIR YRIAKNPGTA TVEDGSLFDI EVGPEG LTF PFVLRYRGHK FPEQLSSVIR YWEENDGKNG MAWLGGLDST GKGRFALKDI KIFEWDLNQK INEYIKERGM RGKEKEL LE MGESSLPDGL IPYKFFEERE CLFPYKENLK PQWSEVQYTI EVGSPLLTAD TISALTEPGN RDAIAYKKRV YNDGNNAI E PEPRFAVKSE THRGIFRTAV GRRTGDLGKE DHEDCTCDMC IIFGNEHESS KIRFEDLELI NGNEFEKLEK HIDHVAIDR FTGGALDKAK FDTYPLAGSP KKPLKLKGRF WIKKGFSGDH KLLITTALSD IRDGLYPLGS KGGVGYGWVA GISIDDNVPD DFKEMINKT EMPLPEEVEE SNNGPINNDY VHPGHQSPKQ DHKNKNIYYP HYFLDSGSKV YREKDIITHE EFTEELLSGK I NCKLETLT PLIIPDTSDE NGLKLQGNKP GHKNYKFFNI NGELMIPGSE LRGMLRTHFE ALTKSCFAIF GEDSTLSWRM NA DEKDYKI DSNSIRKMES QRNPKYRIPD ELQKELRNSG NGLFNRLYTS ERRFWSDVSN KFENSIDYKR EILRCAGRPK NYK GGIIRQ RKDSLMAEEL KVHRLPLYDN FDIPDSAYKA NDHCRKSATC STSRGCRERF TCGIKVRDKN RVFLNAANNN RQYL NNIKK SNHDLYLQYL KGEKKIRFNS KVITGSERSP IDVIAELNER GRQTGFIKLS GLNNSNKSQG NTGTTFNSGW DRFEL NILL DDLETRPSKS DYPRPRLLFT KDQYEYNITK RCERVFEIDK GNKTGYPVDD QIKKNYEDIL DSYDGIKDQE VAERFD TFT RGSKLKVGDL VYFHIDGDNK IDSLIPVRIS RKCASKTLGG KLDKALHPCT GLSDGLCPGC HLFGTTDYKG RVKFGFA KY ENGPEWLITR GNNPERSLTL GVLESPRPAF SIPDDESEIP GRKFYLHHNG WRIIRQKQLE IRETVQPERN VTTEVMDK G NVFSFDVRFE NLREWELGLL LQSLDPGKNI AHKLGKGKPY GFGSVKIKID SLHTFKINSN NDKIKRVPQS DIREYINKG YQKLIEWSGN NSIQKGNVLP QWHVIPHIDK LYKLLWVPFL NDSKLEPDVR YPVLNEESKG YIEGSDYTYK KLGDKDNLPY KTRVKGLTT PWSPWNPFQV IAEHEEQEVN VTGSRPSVTD KIERDGKMV UniProtKB: RAMP superfamily protein |
-Macromolecule #2: crRNA
Macromolecule | Name: crRNA / type: rna / ID: 2 / Number of copies: 1 |
---|---|
Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 35.43209 KDa |
Sequence | String: GUUAUGAAAC AAGAGAAGGA CUUAAUGUCA CGGUACCCAA UUUUCUGCCC CGGACUCCAC GGCUGUUACU AGAGGUUAUG AAACAAGAG AAGGACUUAA UGUCACGGUA C |
-Macromolecule #3: Non-self RNA target
Macromolecule | Name: Non-self RNA target / type: rna / ID: 3 / Number of copies: 1 |
---|---|
Source (natural) | Organism: ![]() |
Molecular weight | Theoretical: 18.097797 KDa |
Sequence | String: CUCUAGUAAC AGCCGUGGAG UCCGGGGCAG AAAAUUGGCA UGGCACUGUA AUUCAG |
-Macromolecule #4: ZINC ION
Macromolecule | Name: ZINC ION / type: ligand / ID: 4 / Number of copies: 4 / Formula: ZN |
---|---|
Molecular weight | Theoretical: 65.409 Da |
-Macromolecule #5: MAGNESIUM ION
Macromolecule | Name: MAGNESIUM ION / type: ligand / ID: 5 / Number of copies: 1 / Formula: MG |
---|---|
Molecular weight | Theoretical: 24.305 Da |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
![]() | single particle reconstruction |
Aggregation state | particle |
-
Sample preparation
Buffer | pH: 8 |
---|---|
Vitrification | Cryogen name: ETHANE |
-
Electron microscopy
Microscope | FEI TITAN KRIOS |
---|---|
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.5 µm |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
-
Image processing
Startup model | Type of model: INSILICO MODEL |
---|---|
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 2.54 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 444936 |
Initial angle assignment | Type: RANDOM ASSIGNMENT |
Final angle assignment | Type: RANDOM ASSIGNMENT |