Extended Data Figure 6. Influence of PAM-1 base-pair composition on Cascade binding affinity and Cas3 cleavage efficiency.

a, Inosine substitution of G_NT-1 led to a mild 2-fold binding defect, suggesting the N2 amine of G_NT-1 is a minor determinant of specificity. b, Binding Kd of E. coli Cascade for all four base-pair combinations at PAM-1. Kd for 5′-ATG, ATA, ATC, and ATT PAMs were determined to be ~10, 20, 40, and 100 nM, respectively. c, Cas3 cleavage efficiency was governed by Cascade’s affinity for the corresponding PAM-containing target (5′-ATG>ATA≫ATT≈ATC). Cascade concentration above the Kd led to efficient Cas3 cleavage.