8yny

From Proteopedia

(Difference between revisions)

Current revision

Structure of Cas9-sgRNA ribonucleoprotein bound to nucleosome

Structural highlights

8yny is a 13 chain structure with sequence from Homo sapiens and Streptococcus pyogenes. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance.
Method:	Electron Microscopy, Resolution 4.52Å
Resources:	FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT

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

The CRISPR-associated endonuclease Cas9 derived from prokaryotes is used as a genome editing, which targets specific genomic loci by single guide RNAs (sgRNAs). The eukaryotes, the target of genome editing, store their genome DNA in chromatin, in which the nucleosome is a basic unit. Despite previous structural analyses focusing on Cas9 cleaving free DNA, structural insights into Cas9 targeting of DNA within nucleosomes are limited, leading to uncertainties in understanding how Cas9 operates in the eukaryotic genome. In the present study, we perform native-polyacrylamide gel electrophoresis (PAGE) analyses and find that Cas9 targets the linker DNA and the entry-exit DNA region of the nucleosome but not the DNA tightly wrapped around the histone octamer. We further determine cryo-electron microscopy (cryo-EM) structure of the Cas9-sgRNA-nucleosome ternary complex that targets linker DNA in nucleosomes. The structure suggests interactions between Cas9 and nucleosomes at multiple sites. Mutants that reduce the interaction between nucleosomal DNA and Cas9 improve nucleosomal DNA cleavage activity in vitro, although inhibition by the interaction between Cas9 and nucleosomes is limited in vivo. These findings will contribute to the development of novel genome editing tools in chromatin.

Structural insights into how Cas9 targets nucleosomes.,Nagamura R, Kujirai T, Kato J, Shuto Y, Kusakizako T, Hirano H, Endo M, Toki S, Saika H, Kurumizaka H, Nureki O Nat Commun. 2024 Dec 30;15(1):10744. doi: 10.1038/s41467-024-54768-z. PMID:39737984^[3]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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
↑ Nagamura R, Kujirai T, Kato J, Shuto Y, Kusakizako T, Hirano H, Endo M, Toki S, Saika H, Kurumizaka H, Nureki O. Structural insights into how Cas9 targets nucleosomes. Nat Commun. 2024 Dec 30;15(1):10744. PMID:39737984 doi:10.1038/s41467-024-54768-z

1 Structural highlights
2 Function
3 Publication Abstract from PubMed
4 References

Proteopedia Page Contributors and Editors (what is this?)

OCA

Retrieved from "http://52.214.119.220/wiki/index.php/8yny"

@@ Line 1: / Line 1: @@
-'''Unreleased structure'''
-The entry 8yny is ON HOLD
+==Structure of Cas9-sgRNA ribonucleoprotein bound to nucleosome==
+<StructureSection load='8yny' size='340' side='right'caption='[[8yny]], [[Resolution|resolution]] 4.52&Aring;' scene=''>
+== Structural highlights ==
+<table><tr><td colspan='2'>[[8yny]] is a 13 chain structure with sequence from [https://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens] and [https://en.wikipedia.org/wiki/Streptococcus_pyogenes Streptococcus pyogenes]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=8YNY OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=8YNY FirstGlance]. <br>
+</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">Electron Microscopy, [[Resolution|Resolution]] 4.52&#8491;</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=8yny FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=8yny OCA], [https://pdbe.org/8yny PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=8yny RCSB], [https://www.ebi.ac.uk/pdbsum/8yny PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=8yny ProSAT]</span></td></tr>
+</table>
+== Function ==
+[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;">
+== Publication Abstract from PubMed ==
+The CRISPR-associated endonuclease Cas9 derived from prokaryotes is used as a genome editing, which targets specific genomic loci by single guide RNAs (sgRNAs). The eukaryotes, the target of genome editing, store their genome DNA in chromatin, in which the nucleosome is a basic unit. Despite previous structural analyses focusing on Cas9 cleaving free DNA, structural insights into Cas9 targeting of DNA within nucleosomes are limited, leading to uncertainties in understanding how Cas9 operates in the eukaryotic genome. In the present study, we perform native-polyacrylamide gel electrophoresis (PAGE) analyses and find that Cas9 targets the linker DNA and the entry-exit DNA region of the nucleosome but not the DNA tightly wrapped around the histone octamer. We further determine cryo-electron microscopy (cryo-EM) structure of the Cas9-sgRNA-nucleosome ternary complex that targets linker DNA in nucleosomes. The structure suggests interactions between Cas9 and nucleosomes at multiple sites. Mutants that reduce the interaction between nucleosomal DNA and Cas9 improve nucleosomal DNA cleavage activity in vitro, although inhibition by the interaction between Cas9 and nucleosomes is limited in vivo. These findings will contribute to the development of novel genome editing tools in chromatin.
-Authors:
+Structural insights into how Cas9 targets nucleosomes.,Nagamura R, Kujirai T, Kato J, Shuto Y, Kusakizako T, Hirano H, Endo M, Toki S, Saika H, Kurumizaka H, Nureki O Nat Commun. 2024 Dec 30;15(1):10744. doi: 10.1038/s41467-024-54768-z. PMID:39737984<ref>PMID:39737984</ref>
-Description:
+From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
-[[Category: Unreleased Structures]]
+</div>
+<div class="pdbe-citations 8yny" style="background-color:#fffaf0;"></div>
+== References ==
+<references/>
+__TOC__
+</StructureSection>
+[[Category: Homo sapiens]]
+[[Category: Large Structures]]
+[[Category: Streptococcus pyogenes]]
+[[Category: Hirano H]]
+[[Category: Kujirai T]]
+[[Category: Kurumizaka H]]
+[[Category: Kusakizako T]]
+[[Category: Nagamura R]]
+[[Category: Nureki O]]

8yny

From Proteopedia

Current revision

Structure of Cas9-sgRNA ribonucleoprotein bound to nucleosome

Structural highlights

Function

Publication Abstract from PubMed

References

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Proteopedia Page Contributors and Editors (what is this?)

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