Nature structural & molecular biology, 2014-09, Vol.21 (9), p.771-777
2014
Volltextzugriff (PDF)
Details
- Autor(en) / Beteiligte
- Titel
- Structures of CRISPR Cas3 offer mechanistic insights into Cascade-activated DNA unwinding and degradation
- Ist Teil von
- Nature structural & molecular biology, 2014-09, Vol.21 (9), p.771-777
- Ort / Verlag
- United States
- Erscheinungsjahr
- 2014
- Quelle
- MEDLINE
- Beschreibungen/Notizen
- CRISPR drives prokaryotic adaptation to invasive nucleic acids such as phages and plasmids, using an RNA-mediated interference mechanism. Interference in type I CRISPR-Cas systems requires a targeting Cascade complex and a degradation machine, Cas3, which contains both nuclease and helicase activities. Here we report the crystal structures of Thermobifida fusca Cas3 bound to single-stranded (ss) DNA substrate and show that it is an obligate 3'-to-5' ssDNase that preferentially accepts substrate directly from the helicase moiety. Conserved residues in the HD-type nuclease coordinate two irons for ssDNA cleavage. We demonstrate ATP coordination and conformational flexibility of the SF2-type helicase domain. Cas3 is specifically guided toward Cascade-bound target DNA by a PAM sequence, through physical interactions with both the nontarget substrate strand and the CasA protein. The sequence of recognition events ensures well-controlled DNA targeting and degradation of foreign DNA by Cascade and Cas3.
- Sprache
- Englisch
- Identifikatoren
- ISSN: 1545-9993eISSN: 1545-9985DOI: 10.1038/nsmb.2875Titel-ID: cdi_pubmed_primary_25132177
- Format
- –
- Schlagworte
- Actinomycetales - chemistry, Actinomycetales - enzymology, Actinomycetales - metabolism, Adenosine Triphosphate - metabolism, Base Sequence, CRISPR-Associated Proteins - chemistry, CRISPR-Associated Proteins - metabolism, Crystallography, X-Ray, DNA Helicases - chemistry, DNA Helicases - metabolism, DNA, Single-Stranded - chemistry, DNA, Single-Stranded - metabolism, Models, Molecular, Molecular Sequence Data, Protein Conformation