Table 1.
Metal ion(s) | Kd (nM) | Average endpoint | |
---|---|---|---|
Transition Me2+ | Other | ||
— | — | 10 000 ± 3000 | 0.66 |
20 mM Zn2+ | — | ≥20 000a | n.d.a |
20 mM Cd2+ | — | 2100 ± 400 | 0.75 |
— | 5 mM Sr2+ | 240 ± 15 | 0.93 |
20 mM Zn2+ | 5 mM Sr2+ | 700 ± 300 | 0.59 |
— | 80 mM Sr2+ | 4 ± 0.5 | 0.98 |
20 mM Zn2+ | 80 mM Sr2+ | 9 ± 0.5 | 0.92 |
20 mM Zn2+ | 5 mM | 2486 ± 400 | 0.55 |
— | 20 mM | 751 ± 200 | 0.75 |
20 mM Zn2+ | 20 mM | 350 ± 120 | 0.65 |
— | 80 mM | 79 ± 35 | 0.26 |
20 mM Zn2+ | 80 mM | 53 ± 6 | 0.28 |
Spin column assay for the determination of Kd values were performed at pH 6.0 and 1 M NH4OAc using trace amounts of 5′-endlabeled ptRNAGly; individual Kd values are based on three to six independent experiments and were calculated by non-linear regression analysis (program Grafit, Erithacus Software) using the equation: fc = ft · [P RNA]free/(Kd + [P RNA]free), where fc = fraction of ptRNA in the complex, and ft = maximum fraction of ptRNA that is able to bind to P RNA (endpoint). Endpoints in the right column are the theoretical ones obtained by the fitting procedure; however, theoretical and experimentally measured endpoints were generally in good agreement; average endpoints were normalized to that for 40 mM Sr2+ (see Figure 3A).
aKd and endpoint values (n.d. = not determined) could not be determined with reasonably low errors owing to very low ribozyme–substrate affinity; the Kd of 20 000 nM in the presence of 20 mM Zn2+ alone is a lower limit estimate.