Table 1.
Effect of base identity on cleavage rate and binding affinity
| Oligonucleotide | k2, min−1 | krel | Kd, nM | Kdrel |
|---|---|---|---|---|
| P1 alone | 1 × 10−6* | (1) | 10 | (1) |
| P1-G | 0.005 | 10,000 | 7 | 0.7 |
| P1-A | 1 × 10−6 | 1 | — | — |
| P1-U | 5 × 10−6 | 5 | — | — |
| P1-C | 1 × 10−6 | 1 | 10 | 1 |
| P2 alone | 1 × 10−6* | (1) | 6 | (1) |
| C-P2 | 1 × 10−6 | 1 | — | — |
| UC-P2 | 2 × 10−6 | 2 | 4.5 | 0.8 |
| GUC-P2 | 0.02 | 20,000 | 6 | 1 |
| AUC-P2† | ≈1 × 10−6 | 1 | — | — |
| UUC-P2 | 0.5 × 10−6 | 0.5 | — | — |
| CUC-P2 | 0.3 × 10−6 | 0.3 | — | — |
Conditions: 10 mM MgCl2, 25°C.
*
These oligonucleotides do not contain the cleavage site; the nonenzymatic cleavage rate, determined as described in Methods, is listed. These values are within 2-fold of those obtained for S, and this small difference in background cleavage rates has no effect on the conclusions drawn from these data.
†
The oligonucleotide AUC-P2 gave a predominant cleavage band after position 16.3, consistent with binding of a fraction of the oligonucleotide in an alternative conformation (with the 5′-terminal A pairing with U15.5 of the ribozyme). The rate constant in the table corresponds to the ≈1% of the reaction that occurs at the normal cleavage position.