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TitleCasX enzymes comprise a distinct family of RNA-guided genome editors.
Journal, issue, pagesNature, Vol. 566, Issue 7743, Page 218-223, Year 2019
Publish dateFeb 4, 2019
AuthorsJun-Jie Liu / Natalia Orlova / Benjamin L Oakes / Enbo Ma / Hannah B Spinner / Katherine L M Baney / Jonathan Chuck / Dan Tan / Gavin J Knott / Lucas B Harrington / Basem Al-Shayeb / Alexander Wagner / Julian Brötzmann / Brett T Staahl / Kian L Taylor / John Desmarais / Eva Nogales / Jennifer A Doudna /
PubMed AbstractThe RNA-guided CRISPR-associated (Cas) proteins Cas9 and Cas12a provide adaptive immunity against invading nucleic acids, and function as powerful tools for genome editing in a wide range of ...The RNA-guided CRISPR-associated (Cas) proteins Cas9 and Cas12a provide adaptive immunity against invading nucleic acids, and function as powerful tools for genome editing in a wide range of organisms. Here we reveal the underlying mechanisms of a third, fundamentally distinct RNA-guided genome-editing platform named CRISPR-CasX, which uses unique structures for programmable double-stranded DNA binding and cleavage. Biochemical and in vivo data demonstrate that CasX is active for Escherichia coli and human genome modification. Eight cryo-electron microscopy structures of CasX in different states of assembly with its guide RNA and double-stranded DNA substrates reveal an extensive RNA scaffold and a domain required for DNA unwinding. These data demonstrate how CasX activity arose through convergent evolution to establish an enzyme family that is functionally separate from both Cas9 and Cas12a.
External linksNature / PubMed:30718774 / PubMed Central
MethodsEM (single particle)
Resolution3.2 - 20.0 Å
Structure data

8980

6ny3

EMDB-8980, PDB-6ny3:
CasX ternary complex with 30bp target DNA
Method: EM (single particle) / Resolution: 3.7 Å

EMDB-8987:
ApoCasX
Method: EM (single particle) / Resolution: 20.0 Å

EMDB-8988:
CasX binary complex
Method: EM (single particle) / Resolution: 7.5 Å

EMDB-8989:
CasX-gRNA-DNA (45T-20NT) State I
Method: EM (single particle) / Resolution: 4.5 Å

EMDB-8990:
CasX-gRNA-DNA (45T-20NT) State II
Method: EM (single particle) / Resolution: 4.4 Å

EMDB-8991:
CasX-gRNA-DNA (45T-45NT) State II
Method: EM (single particle) / Resolution: 5.2 Å

8994

6ny1

EMDB-8994: CasX-gRNA-DNA (30bp) State II
PDB-6ny1: CasX-gRNA-DNA(30bp) State II
Method: EM (single particle) / Resolution: 4.2 Å

8996

6ny2

EMDB-8996, PDB-6ny2:
CasX-gRNA-DNA(45bp) state I
Method: EM (single particle) / Resolution: 3.2 Å

Source
  • Deltaproteobacteria (bacteria)
  • deltaproteobacteria bacterium (bacteria)
  • Deltaproteobacteria bacterium RIFCSPLOWO2_12_FULL_43_16 (bacteria)
KeywordsRNA BINDING PROTEIN/RNA/DNA / CasX / sgRNA / target DNA / CRISPR / RNA BINDING PROTEIN-RNA-DNA complex

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