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Structure paper

TitleSecond Messenger cA Formation within the Composite Csm1 Palm Pocket of Type III-A CRISPR-Cas Csm Complex and Its Release Path.
Journal, issue, pagesMol Cell, Vol. 75, Issue 5, Page 933-943.e6, Year 2019
Publish dateSep 5, 2019
AuthorsNing Jia / Roger Jones / George Sukenick / Dinshaw J Patel /
PubMed AbstractTarget RNA binding to crRNA-bound type III-A CRISPR-Cas multi-subunit Csm surveillance complexes activates cyclic-oligoadenylate (cA) formation from ATP subunits positioned within the composite pair ...Target RNA binding to crRNA-bound type III-A CRISPR-Cas multi-subunit Csm surveillance complexes activates cyclic-oligoadenylate (cA) formation from ATP subunits positioned within the composite pair of Palm domain pockets of the Csm1 subunit. The generated cA second messenger in turn targets the CARF domain of trans-acting RNase Csm6, triggering its HEPN domain-based RNase activity. We have undertaken cryo-EM studies on multi-subunit Thermococcus onnurineus Csm effector ternary complexes, as well as X-ray studies on Csm1-Csm4 cassette, both bound to substrate (AMPPNP), intermediates (pppA), and products (cA), to decipher mechanistic aspects of cA formation and release. A network of intermolecular hydrogen bond alignments accounts for the observed adenosine specificity, with ligand positioning dictating formation of linear pppA intermediates and subsequent cA formation by cyclization. We combine our structural results with published functional studies to highlight mechanistic insights into the role of the Csm effector complex in mediating the cA signaling pathway.
External linksMol Cell / PubMed:31326272 / PubMed Central
MethodsEM (single particle) / X-ray diffraction
Resolution2.4 - 3.2 Å
Structure data

0640

6o7e

EMDB-0640, PDB-6o7e:
Cryo-EM structure of Csm-crRNA-target RNA ternary complex in complex with AMPPNP in type III-A CRISPR-Cas system
Method: EM (single particle) / Resolution: 3.2 Å

0641

6o7h

EMDB-0641, PDB-6o7h:
Cryo-EM structure of Csm-crRNA-target RNA ternary complex in complex with cA4 in type III-A CRISPR-Cas system
Method: EM (single particle) / Resolution: 2.9 Å

0642

6o7i

EMDB-0642, PDB-6o7i:
Cryo-EM structure of Csm-crRNA-target RNA ternary bigger complex in complex with cA4 in type III-A CRISPR-Cas system
Method: EM (single particle) / Resolution: 3.2 Å

PDB-6o73:
Crystal structure of apo Csm1-Csm4 cassette
Method: X-RAY DIFFRACTION / Resolution: 3.0 Å

PDB-6o74:
Crystal structure of Csm1-Csm4 cassette in complex with AMPPNP
Method: X-RAY DIFFRACTION / Resolution: 2.71 Å

PDB-6o75:
Crystal structure of Csm1-Csm4 cassette in complex with pppApA
Method: X-RAY DIFFRACTION / Resolution: 2.6 Å

PDB-6o78:
Crystal structure of Csm1-Csm4 cassette in complex with pppApApA
Method: X-RAY DIFFRACTION / Resolution: 2.8 Å

PDB-6o79:
Crystal structure of Csm1-Csm4 cassette in complex with cA3
Method: X-RAY DIFFRACTION / Resolution: 3.0 Å

PDB-6o7b:
Crystal structure of Csm1-Csm4 cassette in complex with cA4
Method: X-RAY DIFFRACTION / Resolution: 2.4 Å

PDB-6o7d:
Crystal structure of Csm1-Csm4 cassette in complex with one ATP
Method: X-RAY DIFFRACTION / Resolution: 2.81 Å

Chemicals

ChemComp-NI:
NICKEL (II) ION / Nickel

ChemComp-ANP:
PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / AMP-PNP, energy-carrying molecule analogue*YM

ChemComp-MN:
Unknown entry

ChemComp-HOH:
WATER / Water

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM / Adenosine triphosphate

ChemComp-ZN:
Unknown entry

ChemComp-G:
GUANOSINE-5'-MONOPHOSPHATE / Guanosine monophosphate

Source
  • thermococcus onnurineus (archaea)
  • thermococcus onnurineus (strain na1) (archaea)
  • thermococcus onnurineus na1 (archaea)
  • synthetic construct (others)
KeywordsIMMUNE SYSTEM / Type III-A CRISPR-Cas system / apo Csm1-Csm4 cassette / aCsm1-Csm4 cassette in complex with AMPPNP / immune system/rna / Csm1-Csm4 cassette in complex with pppApA / immune system-dna complex / immune system-rna complex / Csm1-Csm4 cassette in complex with pppApApA / immune sytem/rna / Csm1-Csm4 cassette in complex with cA3 / immune sytem-rna complex / Csm1-Csm4 cassette in complex with cA4 / Csm1-Csm4 cassette in complex with one ATP / cryo-EM structure / Csm-crRNA-target RNA ternary complex in complex with AMPPNP / Type III CRISPR-Cas systerm / immune system/rna/dna / Csm-crRNA-target RNA ternary complex in complex with CA4 / immune system-rna-dna complex / Csm-crRNA-target RNA ternary bigger complex in complex with CA4

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