Database of articles cited by EMDB/PDB/SASBDB data

Keywords
Structure methods	All X-ray diffraction NMR electron microscopy small angle scattering neutron diffraction fiber diffraction powder diffraction infrared spectroscopy EPR fluorescence transfer theoretical model Hybrid
Author
Journal
IF	>30 >20 >10 >5 all

Title	Unraveling the mechanisms of PAMless DNA interrogation by SpRY-Cas9.
Journal, issue, pages	Nat Commun, Vol. 15, Issue 1, Page 3663, Year 2024
Publish date	Apr 30, 2024
Authors	Grace N Hibshman / Jack P K Bravo / Matthew M Hooper / Tyler L Dangerfield / Hongshan Zhang / Ilya J Finkelstein / Kenneth A Johnson / David W Taylor /
PubMed Abstract	CRISPR-Cas9 is a powerful tool for genome editing, but the strict requirement for an NGG protospacer-adjacent motif (PAM) sequence immediately next to the DNA target limits the number of editable ...CRISPR-Cas9 is a powerful tool for genome editing, but the strict requirement for an NGG protospacer-adjacent motif (PAM) sequence immediately next to the DNA target limits the number of editable genes. Recently developed Cas9 variants have been engineered with relaxed PAM requirements, including SpG-Cas9 (SpG) and the nearly PAM-less SpRY-Cas9 (SpRY). However, the molecular mechanisms of how SpRY recognizes all potential PAM sequences remains unclear. Here, we combine structural and biochemical approaches to determine how SpRY interrogates DNA and recognizes target sites. Divergent PAM sequences can be accommodated through conformational flexibility within the PAM-interacting region, which facilitates tight binding to off-target DNA sequences. Nuclease activation occurs ~1000-fold slower than for Streptococcus pyogenes Cas9, enabling us to directly visualize multiple on-pathway intermediate states. Experiments with SpG position it as an intermediate enzyme between Cas9 and SpRY. Our findings shed light on the molecular mechanisms of PAMless genome editing.
External links	Nat Commun / PubMed:38688943 / PubMed Central
Methods	EM (single particle)
Resolution	2.79 - 3.69 Å
Structure data	40681 8spq EMDB-40681, PDB-8spq: SpRY-Cas9:gRNA complex targeting TGG PAM DNA Method: EM (single particle) / Resolution: 3.26 Å 40705 8sqh EMDB-40705, PDB-8sqh: SpRY-Cas9:gRNA complex targeting TTC PAM DNA Method: EM (single particle) / Resolution: 3.69 Å 40740 8srs EMDB-40740, PDB-8srs: SpRY-Cas9:gRNA complex targeting TAC PAM DNA Method: EM (single particle) / Resolution: 2.79 Å 41073 8t6o EMDB-41073, PDB-8t6o: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 0 bp R-loop Method: EM (single particle) / Resolution: 3.1 Å 41074 8t6p EMDB-41074, PDB-8t6p: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 2 bp R-loop Method: EM (single particle) / Resolution: 3.15 Å 41079 8t6s EMDB-41079, PDB-8t6s: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 10 bp R-loop Method: EM (single particle) / Resolution: 3.28 Å 41080 8t6t EMDB-41080, PDB-8t6t: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 13 bp R-loop Method: EM (single particle) / Resolution: 3.04 Å 41083 8t6x EMDB-41083, PDB-8t6x: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 18 bp R-loop Method: EM (single particle) / Resolution: 3.08 Å 41084 8t6y EMDB-41084, PDB-8t6y: SpRY-Cas9:gRNA complex bound to non-target DNA with 1 bp R-loop Method: EM (single particle) / Resolution: 2.88 Å 41085 8t76 EMDB-41085, PDB-8t76: SpRY-Cas9:gRNA complex targeting TAC PAM DNA with 3 bp R-loop Method: EM (single particle) / Resolution: 3.42 Å 41086 8t77 EMDB-41086, PDB-8t77: SpRY-Cas9:gRNA complex bound to non-target DNA with 6 bp R-loop Method: EM (single particle) / Resolution: 3.25 Å 41087 8t78 EMDB-41087, PDB-8t78: SpRY-Cas9:gRNA complex bound to non-target DNA with 8 bp R-loop Method: EM (single particle) / Resolution: 3.02 Å 41088 8t79 EMDB-41088, PDB-8t79: SpRY-Cas9:gRNA complex bound to non-target DNA with 10 bp R-loop Method: EM (single particle) / Resolution: 3.04 Å 41093 8t7s EMDB-41093, PDB-8t7s: SpRYmer bound to NAC PAM DNA Method: EM (single particle) / Resolution: 3.01 Å 41775 8tzz EMDB-41775, PDB-8tzz: SpG Cas9 with NGC PAM DNA target Method: EM (single particle) / Resolution: 3.56 Å 41867 8u3y EMDB-41867, PDB-8u3y: SpG Cas9 with NGG PAM DNA target Method: EM (single particle) / Resolution: 3.1 Å
Chemicals	ChemComp-MG: Unknown entry ChemComp-HOH: WATER
Source	streptococcus pyogenes (bacteria) streptococcus pyogenes serotype m1 (bacteria) escherichia phage lambda (virus) synthetic construct (others)
Keywords	IMMUNE SYSTEM / SpRY-Cas9 / CRISPR / Cas9 / IMMUNE SYSTEM/HYDROLASE / IMMUNE SYSTEM-HYDROLASE complex / PAM / R-loop / dimer / IMMUNE SYSTEM/RNA/DNA / SpG-Cas9 / IMMUNE SYSTEM-RNA-DNA complex / IMMUNE SYSTEM/DNA/RNA / IMMUNE SYSTEM-DNA-RNA complex