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| Title | Structural basis for RNA-guided DNA cleavage by IscB-ωRNA and mechanistic comparison with Cas9. |
|---|---|
| Journal, issue, pages | Science, Vol. 376, Issue 6600, Page 1476-1481, Year 2022 |
| Publish date | Jun 24, 2022 |
 Authors | Gabriel Schuler / Chunyi Hu / Ailong Ke /  |
| PubMed Abstract | Class 2 CRISPR effectors Cas9 and Cas12 may have evolved from nucleases in IS200/IS605 transposons. IscB is about two-fifths the size of Cas9 but shares a similar domain organization. The associated ...Class 2 CRISPR effectors Cas9 and Cas12 may have evolved from nucleases in IS200/IS605 transposons. IscB is about two-fifths the size of Cas9 but shares a similar domain organization. The associated ωRNA plays the combined role of CRISPR RNA (crRNA) and trans-activating CRISPR RNA tracrRNA) to guide double-stranded DNA (dsDNA) cleavage. Here we report a 2.78-angstrom cryo-electron microscopy structure of IscB-ωRNA bound to a dsDNA target, revealing the architectural and mechanistic similarities between IscB and Cas9 ribonucleoproteins. Target-adjacent motif recognition, R-loop formation, and DNA cleavage mechanisms are explained at high resolution. ωRNA plays the equivalent function of REC domains in Cas9 and contacts the RNA-DNA heteroduplex. The IscB-specific PLMP domain is dispensable for RNA-guided DNA cleavage. The transition from ancestral IscB to Cas9 involved dwarfing the ωRNA and introducing protein domain replacements. |
 External links | Science / PubMed:35617371 / PubMed Central |
| Methods | EM (single particle) |
| Resolution | 2.74 - 3.2 Å |
| Structure data | EMDB-26782, PDB-7utn: EMDB-26976, PDB-8csz: EMDB-26994, PDB-8ctl: |
| Chemicals |  ChemComp-MG:  ChemComp-ZN: |
| Source |
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 Keywords | RNA BINDING PROTEIN/RNA/DNA /  CRISPR / IscB / HEARO RNA / omega RNA / RNA BINDING PROTEIN-RNA-DNA complex |
