4w8y
Structure of full length Cmr2 from Pyrococcus furiosus (Manganese bound form)
Structural highlights
FunctionCMR2_PYRFU 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), formerly called psiRNA (prokaryotic silencing) in this organism. Part of the Cmr ribonucleoprotein complex which has divalent cation-dependent endoribonuclease activity specific for ssRNA complementary to the crRNA, generating 5' hydroxy- and 3' phosphate or 2'-3' cyclic phosphate termini. It is not known which subunit has endoribonuclease activity. Cmr complex does not cleave ssDNA complementary to the crRNA. Cleavage of invading RNA is guided by the crRNA; substrate cleavage occurs a fixed distance (14 nt) from the 3' end of the crRNA. In vitro reconstitution shows Cmr1-2 and Cmr5 are not necessary for cleavage. Probably not the subunit that cleaves pre-crRNA, as mutation of numerous metal-binding residues have no effect on cleavage by assembled complex.[1] Publication Abstract from PubMedThe Cmr complex is an RNA-guided endonuclease that cleaves foreign RNA targets as part of the CRISPR prokaryotic defense system. We investigated the molecular architecture of the P. furiosus Cmr complex using an integrative structural biology approach. We determined crystal structures of P. furiosus Cmr1, Cmr2, Cmr4, and Cmr6 and combined them with known structural information to interpret the cryo-EM map of the complex. To support structure determination, we obtained residue-specific interaction data using protein crosslinking and mass spectrometry. The resulting pseudoatomic model reveals how the superhelical backbone of the complex is defined by the polymerizing principles of Cmr4 and Cmr5 and how it is capped at the extremities by proteins of similar folds. The inner surface of the superhelix exposes conserved residues of Cmr4 that we show are required for target-cleavage activity. The structural and biochemical data thus identify Cmr4 as the conserved endoribonuclease of the Cmr complex. Structural Model of a CRISPR RNA-Silencing Complex Reveals the RNA-Target Cleavage Activity in Cmr4.,Benda C, Ebert J, Scheltema RA, Schiller HB, Baumgartner M, Bonneau F, Mann M, Conti E Mol Cell. 2014 Oct 2;56(1):43-54. doi: 10.1016/j.molcel.2014.09.002. PMID:25280103[2] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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