Foreign DNA capture during CRISPR-Cas adaptive immunity

Nature. 2015 Nov 26;527(7579):535-8. doi: 10.1038/nature15760. Epub 2015 Oct 21.


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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adaptive Immunity*
  • Bacteriophage M13 / genetics
  • Bacteriophage M13 / immunology
  • Base Sequence
  • CRISPR-Associated Proteins / chemistry
  • CRISPR-Associated Proteins / metabolism*
  • Catalytic Domain
  • Clustered Regularly Interspaced Short Palindromic Repeats / genetics*
  • Crystallography, X-Ray
  • DNA, Viral / chemistry
  • DNA, Viral / genetics*
  • DNA, Viral / immunology*
  • DNA, Viral / metabolism
  • Escherichia coli / enzymology
  • Escherichia coli / genetics
  • Escherichia coli / immunology
  • Escherichia coli / virology
  • Integrases / chemistry
  • Integrases / metabolism
  • Models, Molecular
  • Virus Integration* / genetics
  • Virus Integration* / immunology


  • CRISPR-Associated Proteins
  • DNA, Viral
  • Integrases

Associated data

  • PDB/5DS4
  • PDB/5DS5
  • PDB/5DS6