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A Functional Mini-Integrase in a Two-Protein-type V-C CRISPR System

Wright, Addison V., Wang, Joy Y., Burstein, David, Harrington, Lucas B., Paez-Espino, David, Kyrpides, Nikos C., Iavarone, Anthony T., Banfield, Jillian F., Doudna, Jennifer A.
Molecular cell 2019 v.73 no.4 pp. 727-737.e3
CRISPR sequences, DNA, DNA fragmentation, catalytic activity, gene editing, genes, immunity, models, proteins
CRISPR-Cas immunity requires integration of short, foreign DNA fragments into the host genome at the CRISPR locus, a site consisting of alternating repeat sequences and foreign-derived spacers. In most CRISPR systems, the proteins Cas1 and Cas2 form the integration complex and are both essential for DNA acquisition. Most type V-C and V-D systems lack the cas2 gene and have unusually short CRISPR repeats and spacers. Here, we show that a mini-integrase comprising the type V-C Cas1 protein alone catalyzes DNA integration with a preference for short (17- to 19-base-pair) DNA fragments. The mini-integrase has weak specificity for the CRISPR array. We present evidence that the Cas1 proteins form a tetramer for integration. Our findings support a model of a minimal integrase with an internal ruler mechanism that favors shorter repeats and spacers. This minimal integrase may represent the function of the ancestral Cas1 prior to Cas2 adoption.