Abstract
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
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Adaptive Immunity
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CRISPR-Associated Proteins / chemistry
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CRISPR-Associated Proteins / metabolism
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CRISPR-Associated Proteins / physiology*
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CRISPR-Cas Systems*
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Clustered Regularly Interspaced Short Palindromic Repeats / physiology*
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Crystallography, X-Ray
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Endodeoxyribonucleases / chemistry
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Endodeoxyribonucleases / metabolism
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Endodeoxyribonucleases / physiology*
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Endonucleases / chemistry
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Endonucleases / metabolism
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Endonucleases / physiology*
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Escherichia coli / immunology*
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Escherichia coli / metabolism
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Escherichia coli Proteins / chemistry
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Escherichia coli Proteins / metabolism
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Escherichia coli Proteins / physiology*
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Models, Molecular
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Protein Structure, Tertiary
Substances
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CRISPR-Associated Proteins
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Escherichia coli Proteins
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Cas2 protein, E coli
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Endodeoxyribonucleases
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Endonucleases
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YgbT protein, E coli