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- Title
Foreign DNA capture during CRISPR-Cas adaptive immunity.
- Authors
Nuñez, James K.; Harrington, Lucas B.; Kranzusch, Philip J.; Engelman, Alan N.; Doudna, Jennifer A.
- Abstract
Bacteria and archaea generate adaptive immunity against phages and plasmids by integrating foreign DNA of specific 30-40-base-pair lengths into clustered regularly interspaced short palindromic repeat (CRISPR) loci as spacer segments. The universally conserved Cas1-Cas2 integrase complex catalyses spacer acquisition using a direct nucleophilic integration mechanism similar to retroviral integrases and transposases. How the Cas1-Cas2 complex selects foreign DNA substrates for integration remains unknown. Here we present X-ray crystal structures of the Escherichia coli Cas1-Cas2 complex bound to cognate 33-nucleotide protospacer DNA substrates. The protein complex creates a curved binding surface spanning the length of the DNA and splays the ends of the protospacer to allow each terminal nucleophilic 3′-OH to enter a channel leading into the Cas1 active sites. Phosphodiester backbone interactions between the protospacer and the proteins explain the sequence-nonspecific substrate selection observed in vivo. Our results uncover the structural basis for foreign DNA capture and the mechanism by which Cas1-Cas2 functions as a molecular ruler to dictate the sequence architecture of CRISPR loci.
- Subjects
CRISPRS; LOCUS (Genetics); IMMUNOGENETICS; NUCLEOPHILIC addition (Chemistry); X-ray crystallography; ESCHERICHIA coli DNA
- Publication
Nature, 2015, Vol 527, Issue 7579, p535
- ISSN
0028-0836
- Publication type
Article
- DOI
10.1038/nature15760