Table 1. Rate of cleavage (k_app) of pSu1 and pATSer variants at 10 mM Mg²⁺.

Substrate	Cleavage site	kapp (min-1) With 3'-CCA	kapp (min-1) Without 3'-CCA
pSu1(-1C)	C0	0.5±0.01	0.25±0.005
	M-1	0.7±0.01	0.5±0.01
pSu1(-1A)	C0	1.6±0.01	2.5±0.1
	M-1	0.04±0.0004	0.02±0.001
pSu1(-1G)	C0	0.8±0.004	2±0.07
	M-1	0.03±0.001	0.02±0.0004
pSu1(-1U)	C0	1.4±0.08	0.4±0.01
	M-1	0.03±0.003	0.03±0.003
pATSerCG#	C0	0.0002±0.00003	0.0012±0.00001
	M-1	0.0008±0.00005	0.0022±0.0001
pATSerUG#	C0	0.0013±0.00005	0.0034±0.0002
	M-1	0.0002±0.000005	0.00035±0.00005
pATSerCIno#	C0	0.0004±0.000001	0.001±0.00005
	M-1	0.0012±0.00001	0.0019±0.00002
pATSerCU#	C0	0.002±0.00005	ND
	M-1	0.0014±0.00002	ND
pATSerUG##	C0	0.0009±0.0001	NA
pATSerUGΔ3'AC##	C0	0.0006±0.00004	NA
pATSerUGΔ3'CAC##	C0	0.001±0.0001	NA
pATSerUGΔ3'CCAC##	C0	0.003±0.0008	NA

Each value listed is a mean ± standard deviation determined from three or more independent experiments.

^#C and U correspond to residue identity at the -1 position while G, Ino (inosine) and U refer to residue identity at the discriminator position "+73" (numbering same as in tRNA; Fig 1).

^##k_app values determined at 25 mM Mg²⁺ for these substrates. While these experiments were performed prior to our establishing 10 mM Mg²⁺ as being optimal, the rate and fidelity of cleavage is largely unchanged between 10 to 25 mM Mg²⁺. Δ3'AC, Δ3'CAC and Δ3'CCAC indicates residues in the 3'CCAC motif that were deleted. ND, not determined; NA, not applicable.