View source for 5h9e


==Crystal structure of E. coli Cascade bound to a PAM-containing dsDNA target (32-nt spacer) at 3.20 angstrom resolution.==
<StructureSection load='5h9e' size='340' side='right' caption='[[5h9e]], [[Resolution|resolution]] 3.21&Aring;' scene=''>
== Structural highlights ==
<table><tr><td colspan='2'>[[5h9e]] is a 14 chain structure. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5H9E OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5H9E FirstGlance]. <br>
</td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr>
<tr id='NonStdRes'><td class="sblockLbl"><b>[[Non-Standard_Residue|NonStd Res:]]</b></td><td class="sblockDat"><scene name='pdbligand=23G:GUANOSINE-5-PHOSPHATE-2,3-CYCLIC+PHOSPHATE'>23G</scene></td></tr>
<tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[5h9f|5h9f]]</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=5h9e FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5h9e OCA], [http://pdbe.org/5h9e PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=5h9e RCSB], [http://www.ebi.ac.uk/pdbsum/5h9e PDBsum]</span></td></tr>
</table>
== Function ==
[[http://www.uniprot.org/uniprot/CASC_ECOLI CASC_ECOLI]] 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 sequences complementary to antecedent mobile elements and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA).<ref>PMID:21255106</ref> <ref>PMID:21460843</ref> <ref>PMID:21699496</ref>   A component of Cascade, which participates in CRISPR interference, the third stage of CRISPR immunity. Cascade binds both crRNA and in a sequence-specific manner negatively supercoiled dsDNA target. This leads to the formation of an R-loop in which the crRNA binds the target DNA, displacing the noncomplementary strand. Cas3 is recruited to Cascade, nicks target DNA and then unwinds and cleaves the target, leading to DNA degradation and invader neutralization. CasCDE alone is also able to form R-loops.<ref>PMID:21255106</ref> <ref>PMID:21460843</ref> <ref>PMID:21699496</ref>  [[http://www.uniprot.org/uniprot/CSE2_ECOLI CSE2_ECOLI]] 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 sequences complementary to antecedent mobile elements and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA).<ref>PMID:21255106</ref> <ref>PMID:21460843</ref>   A component of Cascade, which participates in CRISPR interference, the third stage of CRISPR immunity. Cascade binds both crRNA and in a sequence-specific manner negatively supercoiled dsDNA target. This leads to the formation of an R-loop in which the crRNA binds the target DNA, displacing the noncomplementary strand. Cas3 is recruited to Cascade, nicks target DNA and then unwinds and cleaves the target, leading to DNA degradation and invader neutralization.<ref>PMID:21255106</ref> <ref>PMID:21460843</ref>  [[http://www.uniprot.org/uniprot/CAS6_ECOLI CAS6_ECOLI]] 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 sequences complementary to antecedent mobile elements and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA).<ref>PMID:18703739</ref> <ref>PMID:21219465</ref> <ref>PMID:21255106</ref> <ref>PMID:21460843</ref> <ref>PMID:21699496</ref>   CasE is required to process the pre-crRNA into single repeat-spacer units, with an 8-nt 5'-repeat DNA tag that may help other proteins recognize the crRNA. This subunit alone will cleave pre-crRNA, as will CasCDE or CasCE; cleavage does not require divalent metals or ATP. CasCDE alone is also able to form R-loops. Partially inhibits the cleavage of Holliday junctions by YgbT (Cas1). Yields a 5'-hydroxy group and a 2',3'-cyclic phosphate terminus.<ref>PMID:18703739</ref> <ref>PMID:21219465</ref> <ref>PMID:21255106</ref> <ref>PMID:21460843</ref> <ref>PMID:21699496</ref>   A component of Cascade, which participates in CRISPR interference, the third stage of CRISPR immunity. Cascade binds both crRNA and in a sequence-specific manner negatively supercoiled dsDNA target. This leads to the formation of an R-loop in which the crRNA binds the target DNA, displacing the noncomplementary strand. Cas3 is recruited to Cascade, nicks target DNA and then unwinds and cleaves the target, leading to DNA degradation and invader neutralization.<ref>PMID:18703739</ref> <ref>PMID:21219465</ref> <ref>PMID:21255106</ref> <ref>PMID:21460843</ref> <ref>PMID:21699496</ref>  [[http://www.uniprot.org/uniprot/CSE1_ECOLI CSE1_ECOLI]] 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 sequences complementary to antecedent mobile elements and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA).<ref>PMID:21255106</ref> <ref>PMID:21460843</ref> <ref>PMID:22621933</ref> <ref>PMID:22521690</ref>   A component of Cascade, which participates in CRISPR interference, the third stage of CRISPR immunity. Cascade binds both crRNA and in a sequence-specific manner negatively supercoiled dsDNA target. This leads to the formation of an R-loop in which the crRNA binds the target DNA, displacing the noncomplementary strand. Cas3 is recruited to Cascade, probably via interactions with CasA, nicks target DNA and then unwinds and cleaves the target, leading to DNA degradation and invader neutralization. CasA is not required for formation of Cascade, but probably enhances binding to and subsequent recognition of both target dsDNA and ssDNA.<ref>PMID:21255106</ref> <ref>PMID:21460843</ref> <ref>PMID:22621933</ref> <ref>PMID:22521690</ref>  [[http://www.uniprot.org/uniprot/CAS5_ECOLI CAS5_ECOLI]] 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 sequences complementary to antecedent mobile elements and target invading nucleic acids. CRISPR clusters are transcribed and processed into CRISPR RNA (crRNA).<ref>PMID:21255106</ref> <ref>PMID:21460843</ref> <ref>PMID:21699496</ref>   A component of Cascade, which participates in CRISPR interference, the third stage of CRISPR immunity. Cascade binds both crRNA and in a sequence-specific manner negatively supercoiled dsDNA target. This leads to the formation of an R-loop in which the crRNA binds the target DNA, displacing the noncomplementary strand. Cas3 is recruited to Cascade, nicks target DNA and then unwinds and cleaves the target, leading to DNA degradation and invader neutralization. CasCDE alone is also able to form R-loops.<ref>PMID:21255106</ref> <ref>PMID:21460843</ref> <ref>PMID:21699496</ref>  
<div style="background-color:#fffaf0;">
== Publication Abstract from PubMed ==
Clustered regularly interspaced short palindromic repeats (CRISPRs) and the cas (CRISPR-associated) operon form an RNA-based adaptive immune system against foreign genetic elements in prokaryotes. Type I accounts for 95% of CRISPR systems, and has been used to control gene expression and cell fate. During CRISPR RNA (crRNA)-guided interference, Cascade (CRISPR-associated complex for antiviral defence) facilitates the crRNA-guided invasion of double-stranded DNA for complementary base-pairing with the target DNA strand while displacing the non-target strand, forming an R-loop. Cas3, which has nuclease and helicase activities, is subsequently recruited to degrade two DNA strands. A protospacer adjacent motif (PAM) sequence flanking target DNA is crucial for self versus foreign discrimination. Here we present the 2.45 A crystal structure of Escherichia coli Cascade bound to a foreign double-stranded DNA target. The 5'-ATG PAM is recognized in duplex form, from the minor groove side, by three structural features in the Cascade Cse1 subunit. The promiscuity inherent to minor groove DNA recognition rationalizes the observation that a single Cascade complex can respond to several distinct PAM sequences. Optimal PAM recognition coincides with wedge insertion, initiating directional target DNA strand unwinding to allow segmented base-pairing with crRNA. The non-target strand is guided along a parallel path 25 A apart, and the R-loop structure is further stabilized by locking this strand behind the Cse2 dimer. These observations provide the structural basis for understanding the PAM-dependent directional R-loop formation process.

Structural basis for promiscuous PAM recognition in type I-E Cascade from E. coli.,Hayes RP, Xiao Y, Ding F, van Erp PB, Rajashankar K, Bailey S, Wiedenheft B, Ke A Nature. 2016 Feb 25;530(7591):499-503. doi: 10.1038/nature16995. Epub 2016 Feb, 10. PMID:26863189<ref>PMID:26863189</ref>

From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
</div>
<div class="pdbe-citations 5h9e" style="background-color:#fffaf0;"></div>
== References ==
<references/>
__TOC__
</StructureSection>
[[Category: Bailey, S]]
[[Category: Ding, F]]
[[Category: Erp, P B.G van]]
[[Category: Hayes, R P]]
[[Category: Ke, A]]
[[Category: Rajashankar, K]]
[[Category: Wiedenheft, B]]
[[Category: Xiao, Y]]
[[Category: Crispr cascade]]
[[Category: Immune system-rna complex]]