TABLE 1.

Kinetic parameters of AsCas12a pre-crRNA processing

	kon (M−1 sec−1)	koff (sec−1)b	Kd (pM)	kc (sec−1)
pre-crRNA(−34:+24)	2.9 (±0.2) × 106	1.6 (±0.5) × 10−6	0.56 (±0.18)	2.3 (±0.3) × 10−3
pre-crRNA(−23:+24)	2.6 (±0.2) × 106	3.6 (±0.3) × 10−6	1.4 (±0.2)	2.3 (±0.2) × 10−3
pre-crRNA(−34:0)	6.7 (±0.3) × 105	5.3 (±0.7) × 10−6	7.2 (±1.4)	1.6 (±0.5) × 10−3
pre-crRNA(−23:0)	6.2 (±1.2) × 105	6.7 (±0.5) × 10−6	5.0 (±1.3)	1.8 (±0.15) × 10−3
*crRNA(−18:+24)	5.9 (±0.7) × 106	4.5 (±0.3) × 10−6	0.76 (±0.09)	NA
crRNA(−18:+24)*a	1.4 (±0.3) × 107	7.4 (±0.2) × 10−6	0.53 (±0.12)	NA
crRNA(−18:+24)	ND	2.2 (±0.3) × 10−5c	ND	NA
crRNA(−18:0)a	1.7 (±0.2) × 105	6.7 (±0.2) × 10−5	390 (±50)	NA

^aFor crRNA variants, the asterisk indicates whether the radiolabel is at the 5′ end (*crRNA) or the 3′ end (crRNA*).

^bFor the determination of the pre-cRNA dissociation rate constants, we used a pre-crRNA substrate containing dU at position −19 to prevent Cas12a-mediated cleavage.

^cMeasured using excess pre-crRNA_{(−34: + 24)} in multiple turnover reactions.

(NA) Not applicable, (ND) not determined.