Table I.

P1 duplex unwinding of wild-type CYT-19 and truncated Δ578–626 proteins¹

	CYT-19			Δ578–626
Substrate extension2	2 mM Mg2+	5 mM Mg2+	10 mM Mg2+	2 mM Mg2+	5 mM Mg2+	10 mM Mg2+
None	2.2 × 106	6.9 ± 1.7 × 105	1.7 × 105	1.4 × 106	4.1 ± 1.8 × 105	2.0 × 105
Ribozyme	3.5 ± 1.7 × 107	4.1 ± 1.4 × 107	1.5 ± 0.6 × 107	4.3 ± 1.0 × 106	3.6 ± 1.8 × 106	1.3 ± 0.1 × 106
A20	N.D.3	1.0 × 107	3.0 × 106	N.D.	1.4 × 106	3.1 × 105
P2	4.5 × 107	2.7 ± 0.6 × 107	N.D.	2.8 × 106	9.3 ± 1.0 × 105	N.D.
P9.2	N.D.	6.0 × 106	N.D.	N.D.	9.9 × 105	N.D.

¹All values represent second order rate constants (M⁻¹ min⁻¹). Values with associated uncertainty represent the average and standard deviation of two to four independent determinations, and values without associated uncertainty represent a single determination. All reactions were performed at 25 °C, 50 mM Na-MOPS, pH 7.0, and the indicated concentration of Mg²⁺ (as MgCl₂). Reactions of the wild-type protein in the presence of 10 mM Mg²⁺ have been reported previously (11). The measurements reported here were performed side-by-side with the Δ578–626 protein and gave results that were the same within error as those reported previously except for the A₂₀ extension, which previously gave a value of 9.6 (± 2.0) × 10⁵ M⁻¹ min⁻¹. We do not understand the origin of this 3-fold difference, but it does not affect the conclusions of the current work.

²The substrate designation refers to what RNA structure, if any, was covalently connected to the P1 duplex. “None” represents unwinding of the isolated P1 duplex, formed between the substrate (CCCUCUA₅) and an oligonucleotide that mimics the internal guide sequence of the ribozyme (GGAGGGA). This internal guide sequence strand was extended for the other constructs by addition of A₂₀ or of sequences that can form the P2 or P9.2 helices (11).

³N.D. – Not determined.