Table 1.
In vitro transcription mutational analysis of the B. subtilis metE SAM-I riboswitch
| Tertiary Element | Variant | T50,obs, (μM) | Dynamic Range | Fraction terminated, (−) SAM | Fraction terminated, (+) SAM |
|---|---|---|---|---|---|
| wild type | 1.6 ± 0.4 | 0.26 ± 0.01 | 0.66 ± 0.01 | 0.92 ± 0.01 | |
| Δ16 | 1.9 ± 0.5 | 0.66 ± 0.03 | 0.21 ± 0.02 | 0.87 ± 0.03 | |
| IL3a | U157A | N.D. | N.D. | N.D. | N.D. |
| P3 | P3S | N.D. | N.D. | N.D. | N.D. |
| P3M | 2.8 ± 0.6 | 0.56 ± 0.03 | 0.28 ± 0.03 | 0.84 ± 0.03 | |
| IL3b | A89G | 2.0 ± 0.7 | 0.58 ± 0.03 | 0.16 ± 0.03 | 0.74 ± 0.03 |
| A153C | 9 ± 1 | 0.61 ± 0.02 | 0.23 ± 0.01 | 0.84 ± 0.02 | |
| A153G | 6.6 ± 0.9 | 0.69 ± 0.02 | 0.17 ± 0.01 | 0.86 ± 0.02 | |
| Δ153 | 60 ± 60 | 0.16 ± 0.05 | 0.19 ± 0.01 | 0.35 ± 0.05 | |
| A89G/Δ153 | 90 ± 4 | 0.24 ± 0.04 | 0.20 ± 0.01 | 0.44 ± 0.04 | |
| Kink-turn | A57C | 90 ± 70 | 0.10 ± 0.02 | 0.25 ±0.01 | 0.35 ± 0.02 |
| A73C | 2.2 ± 0.4 | 0.46 ± 0.01 | 0.20 ± 0.01 | 0.66 ± 0.01 | |
| U68C | 2.1 ± 0.3 | 0.65 ± 0.01 | 0.24 ± 0.01 | 0.89 ± 0.01 | |
| Kink-turn–bulge | GAU | 2.2 ± 0.3 | 0.66 ± 0.01 | 0.27 ± 0.01 | 0.93 ± 0.01 |
| GUA | 1.9 ± 0.1 | 0.67 ± 0.01 | 0.22 ± 0.01 | 0.89 ± 0.01 | |
| AAU | 1.5 ± 0.2 | 0.70 ± 0.03 | 0.17 ± 0.01 | 0.87 ± 0.03 | |
| Kink-turn 3b•3n | C55G | 1.7 ± 0.5 | 0.57 ± 0.02 | 0.19 ± 0.02 | 0.76 ± 0.02 |
| C74G | 1.7 ± 0.7 | 0.53 ± 0.04 | 0.20 ± 0.03 | 0.73 ± 0.04 | |
| C74U | 2.1 ± 0.3 | 0.46 ±0.02 | 0.18 ± 0.01 | 0.64 ± 0.02 | |
| C55G/C74A | 4 ± 1 | 0.58 ± 0.03 | 0.26 ± 0.02 | 0.84 ± 0.03 | |
| P2a length | P2+1bp | 60 ± 50 | 0.12 ± 0.02 | 0.17 ± 0.01 | 0.29 ± 0.02 |
| P2Δ1bp | 1.4 ± 0.4 | 0.66 ± 0.03 | 0.22 ± 0.01 | 0.88 ± 0.03 | |
| P2Δ2bp | N.D. | N.D. | N.D. | N.D. | |
| P2Δ3bp | N.D. | N.D. | N.D. | N.D. | |
| Pseudoknot–Composition | G64C/G65C | 70 ± 30 | 0.08 ± 0.01 | 0.23 ± 0.01 | 0.31 ± 0.01 |
| U62A | 2.1 ± 0.3 | 0.68 ± 0.02 | 0.17 ± 0.01 | 0.85 ± 0.02 | |
| U62C/U168G | 1.8 ± 0.2 | 0.57 ± 0.01 | 0.23 ± 0.01 | 0.80 ± 0.01 | |
| U63A | 2.1 ± 0.2 | 0.58 ± 0.01 | 0.21 ± 0.01 | 0.79 ± 0.01 | |
| G64C | 2.1 ± 0.4 | 0.56 ± 0.02 | 0.18 ± 0.01 | 0.74 ± 0.02 | |
| G65C | 60 ± 30 | 0.13 ± 0.02 | 0.20 ± 0.01 | 0.33 ± 0.02 | |
| U63A/G64C | 100 ± 300 | 0.13 ± 0.09 | 0.27 ± 0.02 | 0.40 ± 0.09 | |
| Pseudoknot–base triples | A61C | 2.7 ± 0.3 | 0.56 ± 0.01 | 0.20 ± 0.01 | 0.76 ± 0.01 |
| U164C | 2.2 ± 0.3 | 0.62 ± 0.01 | 0.10 ± 0.01 | 0.72 ± 0.01 | |
| A161C | 14 ± 3 | 0.57 ± 0.03 | 0.27 ± 0.02 | 0.84 ± 0.03 | |
| A162C | 4.5 ± 0.9 | 0.63 ± 0.03 | 0.19 ± 0.01 | 0.82 ± 0.03 | |
| A161C/A162C | 2.2 ± 0.4 | 0.64 ± 0.03 | 0.10 ± 0.02 | 0.74 ± 0.03 | |
| Double Mutants | Δ153 U62A | 80 ± 30 | 0.38 ± 0.05 | 0.23 ± 0.01 | 0.61 ± 0.05 |
| Δ153 P2Δ1bp | 29 ± 5 | 0.60 ± 0.04 | 0.25 ± 0.01 | 0.85 ± 0.04 |
N.D. = not detected