5vzl - Revision history

OCA at 14:13, 13 March 2024

OCA — Wed, 13 Mar 2024 14:13:07 GMT

←Older revision		Revision as of 14:13, 13 March 2024
Line 3:		Line 3:
	<SX load='5vzl' size='340' side='right' viewer='molstar' caption='[[5vzl]], [[Resolution\|resolution]] 3.90Å' scene=''>		<SX load='5vzl' size='340' side='right' viewer='molstar' caption='[[5vzl]], [[Resolution\|resolution]] 3.90Å' scene=''>
	== Structural highlights ==		== Structural highlights ==
-	<table><tr><td colspan='2'>[[5vzl]] is a 3 chain structure with sequence from [~~http://en.wikipedia.org/wiki/ ], [http~~://en.wikipedia.org/wiki/Bacteriophage_sp. Bacteriophage sp.] and [~~http~~://en.wikipedia.org/wiki/~~Streptococcus_pyogenes_(serotype_1_/_m1_gas)~~ Streptococcus pyogenes ~~(serotype 1 / m1 gas)~~]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5VZL OCA]. For a <b>guided tour on the structure components</b> use [~~http~~://proteopedia.org/fgij/fg.htm?mol=5VZL FirstGlance]. <br>	+	<table><tr><td colspan='2'>[[5vzl]] is a 3 chain structure with sequence from [https://en.wikipedia.org/wiki/Bacteriophage_sp. Bacteriophage sp.] and [https://en.wikipedia.org/wiki/Streptococcus_pyogenes_M1_GAS Streptococcus pyogenes M1 GAS]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5VZL OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=5VZL FirstGlance]. <br>
-	</td></tr><tr id='~~NonStdRes~~'><td class="sblockLbl"><b>[[~~Non-Standard_Residue~~\|~~NonStd Res~~:]]</b></td><td class="sblockDat"~~><scene name~~=~~'pdbligand=GTP:GUANOSINE-5-TRIPHOSPHATE'>GTP</scene~~></td></tr>	+	</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models\|Method:]]</b></td><td class="sblockDat" id="methodDat">Electron Microscopy, [[Resolution\|Resolution]] 3.9Å</td></tr>
-	<tr id='~~gene~~'><td class="sblockLbl"><b>[[~~Gene~~\|~~Gene~~:]]</b></td><td class="sblockDat">~~csn1, cas9, ERS445054_00848 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode~~=~~Info&srchmode~~=5~~&id=160490 Streptococcus pyogenes (serotype 1~~ / ~~M1 GAS)])~~</td></tr>	+	<tr id='ligand'><td class="sblockLbl"><b>[[Ligand\|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=GTP:GUANOSINE-5-TRIPHOSPHATE'>GTP</scene></td></tr>
-	<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[~~http~~://proteopedia.org/fgij/fg.htm?mol=5vzl FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5vzl OCA], [~~http~~://pdbe.org/5vzl PDBe], [~~http~~://www.rcsb.org/pdb/explore.do?structureId=5vzl RCSB], [~~http~~://www.ebi.ac.uk/pdbsum/5vzl PDBsum], [~~http~~://prosat.h-its.org/prosat/prosatexe?pdbcode=5vzl 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=5vzl FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5vzl OCA], [https://pdbe.org/5vzl PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=5vzl RCSB], [https://www.ebi.ac.uk/pdbsum/5vzl PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=5vzl 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;">~~	+
-	~~== Publication Abstract from PubMed ==~~	+
-	~~CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 gene editing technology is derived from a microbial adaptive immune system, where bacteriophages are often the intended target~~. ~~Natural inhibitors of CRISPR~~-~~Cas9 enable phages to evade immunity~~ and ~~show promise in controlling Cas9-mediated gene editing in human cells~~. ~~However, the mechanism of CRISPR-~~Cas9 ~~inhibition is not known, and the potential applications for Cas9 inhibitor proteins~~ in ~~mammalian cells have not been fully established. We show that~~ the ~~anti-~~CRISPR ~~protein AcrIIA4 binds only to assembled Cas9-single-guide RNA~~ (~~sgRNA) complexes and not to Cas9 protein alone. A 3.9 A resolution cryo-electron microscopy structure of~~ the ~~Cas9-sgRNA-AcrIIA4 complex revealed that the surface of AcrIIA4 is highly acidic and binds with a 1:1 stoichiometry to a region of Cas9 that normally engages the DNA~~ protospacer adjacent motif. Consistent with this binding mode, order-of-addition experiments showed that AcrIIA4 interferes with DNA recognition but has no effect on preformed Cas9-sgRNA-DNA complexes. Timed delivery of AcrIIA4 into human cells as either protein or expression plasmid allows on-target Cas9-mediated gene editing while reducing off-target edits. These results provide a mechanistic understanding of AcrIIA4 function and demonstrate that inhibitors can modulate the extent and outcomes of Cas9-mediated gene editing.	+
-		+
-	~~Disabling Cas9 by an anti-CRISPR DNA mimic.,Shin J, Jiang F, Liu JJ, Bray NL, Rauch BJ, Baik SH, Nogales E, Bondy-Denomy J, Corn JE, Doudna JA Sci Adv. 2017 Jul 12;3(7~~)~~:e1701620~~. ~~doi: 10.1126/sciadv.1701620. eCollection 2017, Jul. PMID:28706995~~<ref>PMID:~~28706995~~</ref>	+
-		+
-	~~From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.~~<br>	+
-	~~</div>~~	+
-	~~<div class="pdbe-citations 5vzl" style="background-color~~:~~#fffaf0;">~~</~~div~~>	+

	==See Also==		==See Also==
Line 28:		Line 19:
	[[Category: Bacteriophage sp]]		[[Category: Bacteriophage sp]]
	[[Category: Large Structures]]		[[Category: Large Structures]]
-	[[Category: ~~Doudna, J A~~]]	+	[[Category: Streptococcus pyogenes M1 GAS]]
-	[[Category: ~~Jiang, F~~]]	+	[[Category: Doudna JA]]
-	[[Category: ~~Liu, J J~~]]	+	[[Category: Jiang F]]
-	[[Category: ~~Nogales, E~~]]	+	[[Category: Liu JJ]]
-	[[Category: ~~Anti-crispr]]~~	+	[[Category: Nogales E]]
-	~~[[Category: Cas9]]~~	+
-	~~[[Category: Crispr]]~~	+
-	~~[[Category: Cryo-em]]~~	+
-	~~[[Category: Gene editing]]~~	+
-	~~[[Category: Immune system-rna complex]]~~	+
-	~~[[Category: Off-target]]~~	+
-	~~[[Category: On-target~~]]	+

OCA at 22:15, 10 April 2020

OCA — Fri, 10 Apr 2020 22:15:20 GMT

←Older revision		Revision as of 22:15, 10 April 2020
Line 3:		Line 3:
	<SX load='5vzl' size='340' side='right' viewer='molstar' caption='[[5vzl]], [[Resolution\|resolution]] 3.90Å' scene=''>		<SX load='5vzl' size='340' side='right' viewer='molstar' caption='[[5vzl]], [[Resolution\|resolution]] 3.90Å' scene=''>
	== Structural highlights ==		== Structural highlights ==
-	<table><tr><td colspan='2'>[[5vzl]] is a 3 chain structure with sequence from [http://en.wikipedia.org/wiki/ ], [http://en.wikipedia.org/wiki/Bacteriophage_sp. Bacteriophage sp.] and [http://en.wikipedia.org/wiki/Streptococcus_pyogenes_(serotype_1_/_m1_gas) Streptococcus pyogenes (serotype 1 / m1 gas)]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5VZL OCA]. For a <b>guided tour on the structure components</b> use [http://~~oca~~.~~weizmann.ac.il/oca-docs~~/fgij/fg.htm?mol=5VZL FirstGlance]. <br>	+	<table><tr><td colspan='2'>[[5vzl]] is a 3 chain structure with sequence from [http://en.wikipedia.org/wiki/ ], [http://en.wikipedia.org/wiki/Bacteriophage_sp. Bacteriophage sp.] and [http://en.wikipedia.org/wiki/Streptococcus_pyogenes_(serotype_1_/_m1_gas) Streptococcus pyogenes (serotype 1 / m1 gas)]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5VZL OCA]. For a <b>guided tour on the structure components</b> use [http://proteopedia.org/fgij/fg.htm?mol=5VZL FirstGlance]. <br>
	</td></tr><tr id='NonStdRes'><td class="sblockLbl"><b>[[Non-Standard_Residue\|NonStd Res:]]</b></td><td class="sblockDat"><scene name='pdbligand=GTP:GUANOSINE-5-TRIPHOSPHATE'>GTP</scene></td></tr>		</td></tr><tr id='NonStdRes'><td class="sblockLbl"><b>[[Non-Standard_Residue\|NonStd Res:]]</b></td><td class="sblockDat"><scene name='pdbligand=GTP:GUANOSINE-5-TRIPHOSPHATE'>GTP</scene></td></tr>
	<tr id='gene'><td class="sblockLbl"><b>[[Gene\|Gene:]]</b></td><td class="sblockDat">csn1, cas9, ERS445054_00848 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=160490 Streptococcus pyogenes (serotype 1 / M1 GAS)])</td></tr>		<tr id='gene'><td class="sblockLbl"><b>[[Gene\|Gene:]]</b></td><td class="sblockDat">csn1, cas9, ERS445054_00848 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=160490 Streptococcus pyogenes (serotype 1 / M1 GAS)])</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=5vzl FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5vzl OCA], [http://pdbe.org/5vzl PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=5vzl RCSB], [http://www.ebi.ac.uk/pdbsum/5vzl PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=5vzl ProSAT]</span></td></tr>	+	<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://proteopedia.org/fgij/fg.htm?mol=5vzl FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5vzl OCA], [http://pdbe.org/5vzl PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=5vzl RCSB], [http://www.ebi.ac.uk/pdbsum/5vzl PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=5vzl ProSAT]</span></td></tr>
	</table>		</table>
	== Function ==		== Function ==

OCA at 19:54, 6 March 2020

OCA — Fri, 06 Mar 2020 19:54:24 GMT

←Older revision		Revision as of 19:54, 6 March 2020
Line 1:		Line 1:

	==cryo-EM structure of the Cas9-sgRNA-AcrIIA4 anti-CRISPR complex==		==cryo-EM structure of the Cas9-sgRNA-AcrIIA4 anti-CRISPR complex==
-	<~~StructureSection~~ load='5vzl' size='340' side='right'caption='[[5vzl]], [[Resolution\|resolution]] 3.90Å' scene=''>	+	<SX load='5vzl' size='340' side='right' viewer='molstar' caption='[[5vzl]], [[Resolution\|resolution]] 3.90Å' scene=''>
	== Structural highlights ==		== Structural highlights ==
	<table><tr><td colspan='2'>[[5vzl]] is a 3 chain structure with sequence from [http://en.wikipedia.org/wiki/ ], [http://en.wikipedia.org/wiki/Bacteriophage_sp. Bacteriophage sp.] and [http://en.wikipedia.org/wiki/Streptococcus_pyogenes_(serotype_1_/_m1_gas) Streptococcus pyogenes (serotype 1 / m1 gas)]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5VZL OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5VZL FirstGlance]. <br>		<table><tr><td colspan='2'>[[5vzl]] is a 3 chain structure with sequence from [http://en.wikipedia.org/wiki/ ], [http://en.wikipedia.org/wiki/Bacteriophage_sp. Bacteriophage sp.] and [http://en.wikipedia.org/wiki/Streptococcus_pyogenes_(serotype_1_/_m1_gas) Streptococcus pyogenes (serotype 1 / m1 gas)]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5VZL OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5VZL FirstGlance]. <br>
Line 25:		Line 25:
	<references/>		<references/>
	__TOC__		__TOC__
-	</~~StructureSection~~>	+	</SX>
	[[Category: Bacteriophage sp]]		[[Category: Bacteriophage sp]]
	[[Category: Large Structures]]		[[Category: Large Structures]]

OCA at 16:57, 20 November 2019

OCA — Wed, 20 Nov 2019 16:57:48 GMT

←Older revision		Revision as of 16:57, 20 November 2019
Line 19:		Line 19:
	</div>		</div>
	<div class="pdbe-citations 5vzl" style="background-color:#fffaf0;"></div>		<div class="pdbe-citations 5vzl" style="background-color:#fffaf0;"></div>
		+
		+	==See Also==
		+	*[[Endonuclease 3D structures\|Endonuclease 3D structures]]
	== References ==		== References ==
	<references/>		<references/>

OCA at 08:16, 20 March 2019

OCA — Wed, 20 Mar 2019 08:16:00 GMT

←Older revision		Revision as of 08:16, 20 March 2019
Line 1:		Line 1:

	==cryo-EM structure of the Cas9-sgRNA-AcrIIA4 anti-CRISPR complex==		==cryo-EM structure of the Cas9-sgRNA-AcrIIA4 anti-CRISPR complex==
-	<StructureSection load='5vzl' size='340' side='right' caption='[[5vzl]], [[Resolution\|resolution]] 3.90Å' scene=''>	+	<StructureSection load='5vzl' size='340' side='right'caption='[[5vzl]], [[Resolution\|resolution]] 3.90Å' scene=''>
	== Structural highlights ==		== Structural highlights ==
	<table><tr><td colspan='2'>[[5vzl]] is a 3 chain structure with sequence from [http://en.wikipedia.org/wiki/ ], [http://en.wikipedia.org/wiki/Bacteriophage_sp. Bacteriophage sp.] and [http://en.wikipedia.org/wiki/Streptococcus_pyogenes_(serotype_1_/_m1_gas) Streptococcus pyogenes (serotype 1 / m1 gas)]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5VZL OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5VZL FirstGlance]. <br>		<table><tr><td colspan='2'>[[5vzl]] is a 3 chain structure with sequence from [http://en.wikipedia.org/wiki/ ], [http://en.wikipedia.org/wiki/Bacteriophage_sp. Bacteriophage sp.] and [http://en.wikipedia.org/wiki/Streptococcus_pyogenes_(serotype_1_/_m1_gas) Streptococcus pyogenes (serotype 1 / m1 gas)]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5VZL OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5VZL FirstGlance]. <br>
Line 24:		Line 24:
	</StructureSection>		</StructureSection>
	[[Category: Bacteriophage sp]]		[[Category: Bacteriophage sp]]
		+	[[Category: Large Structures]]
	[[Category: Doudna, J A]]		[[Category: Doudna, J A]]
	[[Category: Jiang, F]]		[[Category: Jiang, F]]
Line 33:		Line 34:
	[[Category: Cryo-em]]		[[Category: Cryo-em]]
	[[Category: Gene editing]]		[[Category: Gene editing]]
		+	[[Category: Immune system-rna complex]]
	[[Category: Off-target]]		[[Category: Off-target]]
	[[Category: On-target]]		[[Category: On-target]]

OCA: Protected "5vzl" [edit=sysop:move=sysop]

OCA — Thu, 16 Nov 2017 04:55:31 GMT

Protected "5vzl" [edit=sysop:move=sysop]

←Older revision

Revision as of 04:55, 16 November 2017

OCA: New page: ==cryo-EM structure of the Cas9-sgRNA-AcrIIA4 anti-CRISPR complex== == Structural highlights ==
<table><tr><td colspan='2'>[[5vzl]] is a 3 chain structure with sequence from [http://en.wikipedia.org/wiki/ ], [http://en.wikipedia.org/wiki/Bacteriophage_sp. Bacteriophage sp.] and [http://en.wikipedia.org/wiki/Streptococcus_pyogenes_(serotype_1_/_m1_gas) Streptococcus pyogenes (serotype 1 / m1 gas)]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5VZL OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5VZL FirstGlance]. <br>
</td></tr><tr id='NonStdRes'><td class="sblockLbl"><b>[[Non-Standard_Residue|NonStd Res:]]</b></td><td class="sblockDat"><scene name='pdbligand=GTP:GUANOSINE-5-TRIPHOSPHATE'>GTP</scene></td></tr>
<tr id='gene'><td class="sblockLbl"><b>[[Gene|Gene:]]</b></td><td class="sblockDat">csn1, cas9, ERS445054_00848 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=160490 Streptococcus pyogenes (serotype 1 / M1 GAS)])</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=5vzl FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5vzl OCA], [http://pdbe.org/5vzl PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=5vzl RCSB], [http://www.ebi.ac.uk/pdbsum/5vzl PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=5vzl ProSAT]</span></td></tr>
</table>
== 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]
<div style="background-color:#fffaf0;">
== Publication Abstract from PubMed ==
CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 gene editing technology is derived from a microbial adaptive immune system, where bacteriophages are often the intended target. Natural inhibitors of CRISPR-Cas9 enable phages to evade immunity and show promise in controlling Cas9-mediated gene editing in human cells. However, the mechanism of CRISPR-Cas9 inhibition is not known, and the potential applications for Cas9 inhibitor proteins in mammalian cells have not been fully established. We show that the anti-CRISPR protein AcrIIA4 binds only to assembled Cas9-single-guide RNA (sgRNA) complexes and not to Cas9 protein alone. A 3.9 A resolution cryo-electron microscopy structure of the Cas9-sgRNA-AcrIIA4 complex revealed that the surface of AcrIIA4 is highly acidic and binds with a 1:1 stoichiometry to a region of Cas9 that normally engages the DNA protospacer adjacent motif. Consistent with this binding mode, order-of-addition experiments showed that AcrIIA4 interferes with DNA recognition but has no effect on preformed Cas9-sgRNA-DNA complexes. Timed delivery of AcrIIA4 into human cells as either protein or expression plasmid allows on-target Cas9-mediated gene editing while reducing off-target edits. These results provide a mechanistic understanding of AcrIIA4 function and demonstrate that inhibitors can modulate the extent and outcomes of Cas9-mediated gene editing.

Disabling Cas9 by an anti-CRISPR DNA mimic.,Shin J, Jiang F, Liu JJ, Bray NL, Rauch BJ, Baik SH, Nogales E, Bondy-Denomy J, Corn JE, Doudna JA Sci Adv. 2017 Jul 12;3(7):e1701620. doi: 10.1126/sciadv.1701620. eCollection 2017, Jul. PMID:28706995<ref>PMID:28706995</ref>

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br>
</div>
<div class="pdbe-citations 5vzl" style="background-color:#fffaf0;"></div>
== References ==
<references/>
TOC
</StructureSection>
[[Category: Bacteriophage sp]]
[[Category: Doudna, J A]]
[[Category: Jiang, F]]
[[Category: Liu, J J]]
[[Category: Nogales, E]]
[[Category: Anti-crispr]]
[[Category: Cas9]]
[[Category: Crispr]]
[[Category: Cryo-em]]
[[Category: Gene editing]]
[[Category: Off-target]]
[[Category: On-target]]