Kinetic dissection of pre-crRNA binding and processing by CRISPR–Cas12a

Binding and dissociation kinetics for mature crRNA. (A) Binding (left and center) and dissociation (right) kinetics of the mature crRNA_−18:+24 with a 5′ label. In the center plot, binding data are shown as the average and SEM from three independent measurements. Values of k_on and k_off were 5.9 (±0.7) × 10⁶ M⁻¹ sec⁻¹ and 4.5 (±0.3) × 10⁻⁶ sec⁻¹, respectively. (B) Binding (left and center) and dissociation (right) kinetics of the mature crRNA_−18:+24 with a 3′ label. In the center plot, binding data are shown as the average and SEM from three independent measurements. Values of k_on and k_off were 1.4 (±0.3) × 10⁷ M⁻¹ sec⁻¹ and 7.4 (±0.2) × 10⁻⁶ sec⁻¹, respectively. In both panels, the binding time courses are fit by pseudo-first-order rate equations (left), the concentration dependences are fit by a line (middle), and the dissociation time courses are fit by a first-order rate equation (right).