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	Mechanism for Cas4-assisted directional spacer acquisition in CRISPR-Cas.
Journal, issue, pages	Nature, Vol. 598, Issue 7881, Page 515-520, Year 2021
Publish date	Sep 29, 2021
Authors	Chunyi Hu / Cristóbal Almendros / Ki Hyun Nam / Ana Rita Costa / Jochem N A Vink / Anna C Haagsma / Saket R Bagde / Stan J J Brouns / Ailong Ke /
PubMed Abstract	Prokaryotes adapt to challenges from mobile genetic elements by integrating spacers derived from foreign DNA in the CRISPR array. Spacer insertion is carried out by the Cas1-Cas2 integrase complex. A ...Prokaryotes adapt to challenges from mobile genetic elements by integrating spacers derived from foreign DNA in the CRISPR array. Spacer insertion is carried out by the Cas1-Cas2 integrase complex. A substantial fraction of CRISPR-Cas systems use a Fe-S cluster containing Cas4 nuclease to ensure that spacers are acquired from DNA flanked by a protospacer adjacent motif (PAM) and inserted into the CRISPR array unidirectionally, so that the transcribed CRISPR RNA can guide target searching in a PAM-dependent manner. Here we provide a high-resolution mechanistic explanation for the Cas4-assisted PAM selection, spacer biogenesis and directional integration by type I-G CRISPR in Geobacter sulfurreducens, in which Cas4 is naturally fused with Cas1, forming Cas4/Cas1. During biogenesis, only DNA duplexes possessing a PAM-embedded 3'-overhang trigger Cas4/Cas1-Cas2 assembly. During this process, the PAM overhang is specifically recognized and sequestered, but is not cleaved by Cas4. This 'molecular constipation' prevents the PAM-side prespacer from participating in integration. Lacking such sequestration, the non-PAM overhang is trimmed by host nucleases and integrated to the leader-side CRISPR repeat. Half-integration subsequently triggers PAM cleavage and Cas4 dissociation, allowing spacer-side integration. Overall, the intricate molecular interaction between Cas4 and Cas1-Cas2 selects PAM-containing prespacers for integration and couples the timing of PAM processing with the stepwise integration to establish directionality.
External links	Nature / PubMed:34588691 / PubMed Central
Methods	EM (single particle)
Resolution	3.2 - 5.8 Å
Structure data	23839 7mi4 EMDB-23839, PDB-7mi4: Symmetrical PAM-PAM prespacer bound Cas4/Cas1/Cas2 complex Method: EM (single particle) / Resolution: 3.2 Å 23840 7mi5 EMDB-23840, PDB-7mi5: Asymmetrical PAM-Non PAM prespacer bound Cas4/Cas1/Cas2 complex Method: EM (single particle) / Resolution: 3.57 Å 23843 7mi9 EMDB-23843, PDB-7mi9: Full integration complex of Cas1/Cas2 from Cas4-containing system Method: EM (single particle) / Resolution: 3.89 Å 23845 7mib EMDB-23845: Half integration complex of Cas4/Cas1/Cas2 and Cas4 still in the Non-PAM side PDB-7mib: Half integration complex of Cas4/Cas1/Cas2 with Cas4 still on the Non-PAM side Method: EM (single particle) / Resolution: 5.8 Å 23847 7mid EMDB-23847, PDB-7mid: Sub-complex of Cas4-Cas1-Cas2 bound PAM containing DNA Method: EM (single particle) / Resolution: 3.56 Å EMDB-23849: Half integration complex of Cas1-Cas2 in Cas4 containing system Method: EM (single particle) / Resolution: 5.8 Å
Chemicals	ChemComp-SF4: IRON/SULFUR CLUSTER / Iron–sulfur cluster ChemComp-MN: Unknown entry
Source	geobacter sulfurreducens (bacteria) Geobacter sulfurreducens
Keywords	HYDROLASE/DNA / CRISPR/Cas / Cas4 / PAM recognition / HYDROLASE-DNA complex / full integration / half integration