Cas1-Cas2 complex formation mediates spacer acquisition during CRISPR-Cas adaptive immunity

Nat Struct Mol Biol. 2014 Jun;21(6):528-34. doi: 10.1038/nsmb.2820. Epub 2014 May 4.


The initial stage of CRISPR-Cas immunity involves the integration of foreign DNA spacer segments into the host genomic CRISPR locus. The nucleases Cas1 and Cas2 are the only proteins conserved among all CRISPR-Cas systems, yet the molecular functions of these proteins during immunity are unknown. Here we show that Cas1 and Cas2 from Escherichia coli form a stable complex that is essential for spacer acquisition and determine the 2.3-Å-resolution crystal structure of the Cas1-Cas2 complex. Mutations that perturb Cas1-Cas2 complex formation disrupt CRISPR DNA recognition and spacer acquisition in vivo. Active site mutants of Cas2, unlike those of Cas1, can still acquire new spacers, thus indicating a nonenzymatic role of Cas2 during immunity. These results reveal the universal roles of Cas1 and Cas2 and suggest a mechanism by which Cas1-Cas2 complexes specify sites of CRISPR spacer integration.

Publication types

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

MeSH terms

  • Adaptive Immunity
  • CRISPR-Associated Proteins / chemistry
  • CRISPR-Associated Proteins / metabolism
  • CRISPR-Associated Proteins / physiology*
  • CRISPR-Cas Systems*
  • Clustered Regularly Interspaced Short Palindromic Repeats / physiology*
  • Crystallography, X-Ray
  • Endodeoxyribonucleases / chemistry
  • Endodeoxyribonucleases / metabolism
  • Endodeoxyribonucleases / physiology*
  • Endonucleases / chemistry
  • Endonucleases / metabolism
  • Endonucleases / physiology*
  • Escherichia coli / immunology*
  • Escherichia coli / metabolism
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / metabolism
  • Escherichia coli Proteins / physiology*
  • Models, Molecular
  • Protein Structure, Tertiary


  • CRISPR-Associated Proteins
  • Escherichia coli Proteins
  • Cas2 protein, E coli
  • Endodeoxyribonucleases
  • Endonucleases
  • YgbT protein, E coli

Associated data

  • PDB/4P6I