5vw1

Crystal structure of SpyCas9-sgRNA-AcrIIA4 ternary complex

spinqualitylabelsall models 5vw1, resolution 2.60Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 5vw1 is a 3 chain structure with sequence from Listeria monocytogenes, Listeria monocytogenes M7 and Streptococcus pyogenes serotype M1. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 2.598Å Ligands: EDO, MG, PEG, PG4 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. Publication Abstract from PubMed Prokaryotic CRISPR-Cas adaptive immune systems utilize sequence-specific RNA-guided endonucleases to defend against infection by viruses, bacteriophages, and mobile elements, while these foreign genetic elements evolve diverse anti-CRISPR proteins to overcome the CRISPR-Cas-mediated defense of the host. Recently, AcrIIA2 and AcrIIA4, encoded by Listeria monocytogene prophages, were shown to block the endonuclease activity of type II-A Streptococcus pyogene Cas9 (SpyCas9). We now report the crystal structure of AcrIIA4 in complex with single-guide RNA-bound SpyCas9, thereby establishing that AcrIIA4 preferentially targets critical residues essential for PAM duplex recognition, as well as blocks target DNA access to key catalytic residues lining the RuvC pocket. These structural insights, validated by biochemical assays on key mutants, demonstrate that AcrIIA4 competitively occupies both PAM-interacting and non-target DNA strand cleavage catalytic pockets. Our studies provide insights into anti-CRISPR-mediated suppression mechanisms for inactivating SpyCas9, thereby broadening the applicability of CRISPR-Cas regulatory tools for genome editing. Inhibition Mechanism of an Anti-CRISPR Suppressor AcrIIA4 Targeting SpyCas9.,Yang H, Patel DJ Mol Cell. 2017 Jul 6;67(1):117-127.e5. doi: 10.1016/j.molcel.2017.05.024. Epub, 2017 Jun 9. PMID:28602637 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See Also Endonuclease 3D structures

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