Extended Data Figure 7: Structural analysis and perturbation of the HNH–RuvC III linker.
From: Conformational control of DNA target cleavage by CRISPR–Cas9
a, Molecules A (left) and B (right) of the sgRNA/DNA-bound Cas9 crystal structure (PDB 4OO8). Molecule A has an ordered HNH domain and HNH–RuvC III linker, whereas these are both disordered in molecule B; the missing density for the HNH domain is replaced with the modelled docked state (right). Another prominent difference is the N-terminal region of the RuvC III motif (blue helices), which rearranges from a helix–loop–helix in molecule A into an extended helix in molecule B. Proline pairs were inserted to prevent formation of this extended helix. b, Target (red) and non-target (black) strand cleavage time courses with the indicated Cas9 variant. Exponential fits are shown as solid lines. c, Kinetics of target (red) and non-target (black) strand cleavage for the indicated Cas9 mutants. ND, cleavage not detected. Error bars in b and c, s.d.; n = 3.
