Structural basis of a Tn7-like transposase recruitment and DNA loading to CRISPR-Cas surveillance complex

摘要

a Schematic diagram of genes involved in transposon-encoded type I-F CRISPR-Cas systems in V. choleraeTn6677. b, c Schematic rendering of arrangement of subunits and ribbon representation of overall structures of Cascade-TniQ complex in pre-target-bound (b) and target dsDNA-bound (c) states. d–f Recognition of TniQ homodimer by subunits Cas6 and Cas7.1 of Cascade. Detailed hydrophilic interactions between TniQ and Cas6 (e) and TniQ and Cas7.1 (f) are shown in the expanded panels. g Mutation analysis of the TniQ residues involved in binding to Cascade and dimerization by His pull-down assay. Unrelated anti-CRISPR protein AcrIIC1 is used as a negative control. h PCR analysis of in vivo transposition with TniQ mutants. The plasmid without Cascade genes is used as a negative control. i PAM recognition by Cas8 subunit. j Elongation of helical pitch of crRNA after DNA loading. k Domain movements occur upon target dsDNA loading. Vector lengths are correlated with the domain motion scale.