Fig. 4: PAM recognition.
From: Structure and engineering of Brevibacillus laterosporus Cas9
a Binding of the PAM duplex to BlCas9. b Schematics of the PAM recognition by BlCas9. Hydrogen bonds are depicted by dashed lines. c and d Recognition of the N4CNAN PAM. Nucleotide T(− 7) and residues Thr1025 and Ala1027 are depicted by space-filling models. Hydrogen bonding and electrostatic interactions are shown as green dashed lines. e In vitro DNA cleavage activities of the wild-type BlCas9 (WT) and PAM recognition mutants. The linearized plasmid targets with the T3CCCAA PAM were incubated with the BlCas9–sgRNA complex at 37 °C for 2 min. Data are mean ± s.d. (n = 3). f In vitro DNA cleavage activities of WT BlCas9, the T1025A variant, and enBlCas9 toward DNA targets with different PAMs. The linearized plasmid targets were incubated at 37 °C for 2 min. Data are mean ± s.d. (n = 3).
