5ng6
Crystal structure of FnCas12a bound to a crRNACrystal structure of FnCas12a bound to a crRNA
Structural highlights
Function[CPF1_FRATN] 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). This protein is a single-RNA guided endonuclease that is also capable of guiding crRNA processing; correct processing of pre-crRNA requires only this protein and the CRISPR locus (PubMed:26422227). When this protein is expressed in E.coli it prevents plasmids homologous to the first CRISPR spacer from transforming, showing it is responsible for plasmid immunity (PubMed:26422227). Has dsDNA endonuclease activity, results in staggered 5-base 5' overhangs 18 and 23 bases downstream of the PAM (protospacer adjacent motif) on the non-targeted and targeted strands respectively (PubMed:26422227).[1] Publication Abstract from PubMedThe CRISPR-associated protein Cas12a (Cpf1), which has been repurposed for genome editing, possesses two distinct nuclease activities: endoribonuclease activity for processing its own guide RNAs and RNA-guided DNase activity for target DNA cleavage. To elucidate the molecular basis of both activities, we determined crystal structures of Francisella novicida Cas12a bound to guide RNA and in complex with an R-loop formed by a non-cleavable guide RNA precursor and a full-length target DNA. Corroborated by biochemical experiments, these structures reveal the mechanisms of guide RNA processing and pre-ordering of the seed sequence in the guide RNA that primes Cas12a for target DNA binding. Furthermore, the R-loop complex structure reveals the strand displacement mechanism that facilitates guide-target hybridization and suggests a mechanism for double-stranded DNA cleavage involving a single active site. Together, these insights advance our mechanistic understanding of Cas12a enzymes and may contribute to further development of genome editing technologies. Structural Basis for Guide RNA Processing and Seed-Dependent DNA Targeting by CRISPR-Cas12a.,Swarts DC, van der Oost J, Jinek M Mol Cell. 2017 Apr 20;66(2):221-233.e4. doi: 10.1016/j.molcel.2017.03.016. PMID:28431230[2] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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