Fig. 2: PAM recognition by Cas10d.
From: Structural rearrangements allow nucleic acid discrimination by type I-D Cascade
a R-loop formation in the type I-D complex. The TS travels up along the Cas7 filament, while the NTS travels across the face of Cas10d. b PAM recognition by Cas10d. A glycine loop from the PAM recognition domain of Cas10d integrates into the minor groove of the PAM in the DNA duplex. c Residues responsible for stabilizing and recognizing PAM nucleotides. Eight residues were found to be within 3.5 Å of the phospho-diester backbone of the dsDNA target. d Alternate angle of the glycine loop locked in the minor groove of the PAM. e Lysine 326 of Cas10d hydrogen bonds with C12 of the target strand PAM and T5 of the non-target strand PAM. These correspond to positions 1 and 2 within the PAM. f Electrophoretic mobility shift assay of fluorescently labeled protospacer dsDNA with a 5′-GTT-3′ PAM incubated with wild-type I-D Cascade, or mutated complexes containing Cas10d(K326A), Cas10d(K326P), Cas10d(∆Loop), or Cas5(K114A). Representative of three independent experiments. Source data are provided as a Source data file.
