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| | ==Nuclease-inactive Streptococcus pyogenes Cas9 (D10A/H840A, dCas9) in complex with single-guide RNA at 3.1 Angstrom resolution== | | ==Nuclease-inactive Streptococcus pyogenes Cas9 (D10A/H840A, dCas9) in complex with single-guide RNA at 3.1 Angstrom resolution== |
| - | <StructureSection load='4zt9' size='340' side='right' caption='[[4zt9]], [[Resolution|resolution]] 3.10Å' scene=''> | + | <StructureSection load='4zt9' size='340' side='right'caption='[[4zt9]], [[Resolution|resolution]] 3.10Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| - | <table><tr><td colspan='2'>[[4zt9]] is a 4 chain structure. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4ZT9 OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4ZT9 FirstGlance]. <br> | + | <table><tr><td colspan='2'>[[4zt9]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Streptococcus_pyogenes Streptococcus pyogenes]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4ZT9 OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=4ZT9 FirstGlance]. <br> |
| - | </td></tr><tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[4zt0|4zt0]]</td></tr> | + | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 3.1Å</td></tr> |
| - | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4zt9 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4zt9 OCA], [http://pdbe.org/4zt9 PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=4zt9 RCSB], [http://www.ebi.ac.uk/pdbsum/4zt9 PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=4zt9 ProSAT]</span></td></tr> | + | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=4zt9 FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4zt9 OCA], [https://pdbe.org/4zt9 PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=4zt9 RCSB], [https://www.ebi.ac.uk/pdbsum/4zt9 PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=4zt9 ProSAT]</span></td></tr> |
| | </table> | | </table> |
| | == Function == | | == Function == |
| - | [[http://www.uniprot.org/uniprot/A0A0C6FZC2_STRPY A0A0C6FZC2_STRPY]] CRISPR (clustered regularly interspaced short palindromic repeat) is an adaptive immune system that provides protection against mobile genetic elements (viruses, transposable elements and conjugative plasmids). CRISPR clusters contain spacers, sequences complementary to antecedent mobile elements, and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA). In type II CRISPR systems correct processing of pre-crRNA requires a trans-encoded small RNA (tracrRNA), endogenous ribonuclease 3 (rnc) and this protein. The tracrRNA serves as a guide for ribonuclease 3-aided processing of pre-crRNA. Subsequently Cas9/crRNA/tracrRNA endonucleolytically cleaves linear or circular dsDNA target complementary to the spacer; Cas9 is inactive in the absence of the 2 guide RNAs (gRNA). Cas9 recognizes the protospacer adjacent motif (PAM) in the CRISPR repeat sequences to help distinguish self versus nonself, as targets within the bacterial CRISPR locus do not have PAMs. PAM recognition is also required for catalytic activity.[HAMAP-Rule:MF_01480] | + | [https://www.uniprot.org/uniprot/CAS9_STRP1 CAS9_STRP1] CRISPR (clustered regularly interspaced short palindromic repeat) is an adaptive immune system that provides protection against mobile genetic elements (viruses, transposable elements and conjugative plasmids). CRISPR clusters contain spacers, sequences complementary to antecedent mobile elements, and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA) (Probable). In type II CRISPR systems correct processing of pre-crRNA requires a trans-encoded small RNA (tracrRNA), endogenous ribonuclease 3 (rnc) and this protein. The tracrRNA serves as a guide for ribonuclease 3-aided processing of pre-crRNA. Subsequently Cas9/crRNA/tracrRNA endonucleolytically cleaves linear or circular dsDNA target complementary to the spacer. The target strand not complementary to crRNA is first cut endonucleolytically, then trimmed by 3'-5' exonucleolytically. DNA-binding requires protein and both RNA species. Cas9 probably recognizes a short motif in the CRISPR repeat sequences (the PAM or protospacer adjacent motif) to help distinguish self versus nonself.<ref>PMID:21455174</ref> <ref>PMID:22745249</ref> |
| | <div style="background-color:#fffaf0;"> | | <div style="background-color:#fffaf0;"> |
| | == Publication Abstract from PubMed == | | == Publication Abstract from PubMed == |
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| | *[[CRISPR-Cas|CRISPR-Cas]] | | *[[CRISPR-Cas|CRISPR-Cas]] |
| | *[[CRISPR-Cas9|CRISPR-Cas9]] | | *[[CRISPR-Cas9|CRISPR-Cas9]] |
| | + | *[[Endonuclease 3D structures|Endonuclease 3D structures]] |
| | == References == | | == References == |
| | <references/> | | <references/> |
| | __TOC__ | | __TOC__ |
| | </StructureSection> | | </StructureSection> |
| - | [[Category: Doudna, J A]] | + | [[Category: Large Structures]] |
| - | [[Category: Jiang, F]] | + | [[Category: Streptococcus pyogenes]] |
| - | [[Category: Bacteria adaptive immunity]] | + | [[Category: Doudna JA]] |
| - | [[Category: Crispr-cas9]] | + | [[Category: Jiang F]] |
| - | [[Category: Genome editing and regulation]]
| + | |
| - | [[Category: Hydrolase-rna complex]]
| + | |
| Structural highlights
Function
CAS9_STRP1 CRISPR (clustered regularly interspaced short palindromic repeat) is an adaptive immune system that provides protection against mobile genetic elements (viruses, transposable elements and conjugative plasmids). CRISPR clusters contain spacers, sequences complementary to antecedent mobile elements, and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA) (Probable). In type II CRISPR systems correct processing of pre-crRNA requires a trans-encoded small RNA (tracrRNA), endogenous ribonuclease 3 (rnc) and this protein. The tracrRNA serves as a guide for ribonuclease 3-aided processing of pre-crRNA. Subsequently Cas9/crRNA/tracrRNA endonucleolytically cleaves linear or circular dsDNA target complementary to the spacer. The target strand not complementary to crRNA is first cut endonucleolytically, then trimmed by 3'-5' exonucleolytically. DNA-binding requires protein and both RNA species. Cas9 probably recognizes a short motif in the CRISPR repeat sequences (the PAM or protospacer adjacent motif) to help distinguish self versus nonself.[1] [2]
Publication Abstract from PubMed
Bacterial adaptive immunity uses CRISPR (clustered regularly interspaced short palindromic repeats)-associated (Cas) proteins together with CRISPR transcripts for foreign DNA degradation. In type II CRISPR-Cas systems, activation of Cas9 endonuclease for DNA recognition upon guide RNA binding occurs by an unknown mechanism. Crystal structures of Cas9 bound to single-guide RNA reveal a conformation distinct from both the apo and DNA-bound states, in which the 10-nucleotide RNA "seed" sequence required for initial DNA interrogation is preordered in an A-form conformation. This segment of the guide RNA is essential for Cas9 to form a DNA recognition-competent structure that is poised to engage double-stranded DNA target sequences. We construe this as convergent evolution of a "seed" mechanism reminiscent of that used by Argonaute proteins during RNA interference in eukaryotes.
STRUCTURAL BIOLOGY. A Cas9-guide RNA complex preorganized for target DNA recognition.,Jiang F, Zhou K, Ma L, Gressel S, Doudna JA Science. 2015 Jun 26;348(6242):1477-81. doi: 10.1126/science.aab1452. PMID:26113724[3]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Deltcheva E, Chylinski K, Sharma CM, Gonzales K, Chao Y, Pirzada ZA, Eckert MR, Vogel J, Charpentier E. CRISPR RNA maturation by trans-encoded small RNA and host factor RNase III. Nature. 2011 Mar 31;471(7340):602-7. doi: 10.1038/nature09886. PMID:21455174 doi:http://dx.doi.org/10.1038/nature09886
- ↑ Jinek M, Chylinski K, Fonfara I, Hauer M, Doudna JA, Charpentier E. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science. 2012 Aug 17;337(6096):816-21. doi: 10.1126/science.1225829. Epub 2012, Jun 28. PMID:22745249 doi:http://dx.doi.org/10.1126/science.1225829
- ↑ Jiang F, Zhou K, Ma L, Gressel S, Doudna JA. STRUCTURAL BIOLOGY. A Cas9-guide RNA complex preorganized for target DNA recognition. Science. 2015 Jun 26;348(6242):1477-81. doi: 10.1126/science.aab1452. PMID:26113724 doi:http://dx.doi.org/10.1126/science.aab1452
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