Table 1.
Binding of wild-type and mutant HsPUM1-HD proteins to wild-type and mutant RNAs
| Protein | RNA | RNA sequence* | KD, nM | Trials | Krel† |
|---|---|---|---|---|---|
| 87654321 | |||||
| WT protein | NRE1–19 (WT RNA) | CCAGAAUUGUAUAUAUUCG | 0.48 ± 0.21 | 6 | 1 |
| WT protein | G2U | CCAGAAUUUUAUAUAUUCG | 59 ± 12 | 3 | 123 |
| WT protein | G2A | CCAGAAUUAUAUAUAUUCG | 16 ± 3.9 | 4 | 33 |
| WT protein | GU23UG | CCAGAAUUUGAUAUAUUCG | 1,600 ± 420 | 3 | 3,300 |
| WT protein | A8G | CCAGAAUUGUAUAUGUUCG | 1.6 ± 0.15 | 3 | 3.3 |
| WT protein | A6G | CCAGAAUUGUAUGUAUUCG | 0.72 ± 0.11 | 3 | 1.5 |
| WT protein | A6U | CCAGAAUUGUAUUUAUUCG | 8.0 ± 3.1 | 3 | 17 |
| WT protein | ARE | UUAUUUAUUUAUUUAUU | 11 ± 3.4 | 4 | 23 |
| G → U: MUT7-2 (S1079N/E1083Q) | |||||
| MUT7-2 | G2U | CCAGAAUUUUAUAUAUUCG | 6.0 ± 1.3 | 3 | 1 |
| MUT7-2 | WT RNA | CCAGAAUUGUAUAUAUUCG | 150 ± 28 | 3 | 25 |
| U → G, G → U: MUT6-2/7-2 (N1043S/Q1047E/S1079N/E1083Q) | |||||
| MUT6-2/7-2 | GU23UG | CCAGAAUUUGAUAUAUUCG | 18 ± 6.7 | 5 | 1 |
| MUT6-2/7-2 | WT RNA | CCAGAAUUGUAUAUAUUCG | 610 ± 78 | 3 | 34 |
| A → G: MUT1-1 (Q867E) | |||||
| MUT1-1 | A8G | CCAGAAUUGUAUAUGUUCG | 1.3 ± 0.38 | 5 | 1 |
| MUT1-1 | WT RNA | CCAGAAUUGUAUAUAUUCG | 190 ± 42 | 4 | 146 |
| A → G: MUT3-2 (C935S/Q939E) | |||||
| MUT3-2 | A6G | CCAGAAUUGUAUGUAUUCG | 0.051 ± 0.004 | 5 | 1 |
| MUT3-2 | WT RNA | CCAGAAUUGUAUAUAUUCG | 2.8 ± 0.36 | 4 | 55 |
| MUT7-1 (E1083Q) | |||||
| MUT7-1 | G2U | CCAGAAUUUUAUAUAUUCG | 5.7 ± 0.55 | 4 | 1 |
| MUT7-1 | G2A | CCAGAAUUAUAUAUAUUCG | 35 ± 5.4 | 3 | 6.1 |
| MUT7-1 | WT RNA | CCAGAAUUGUAUAUAUUCG | 7.7 ± 1.1 | 3 | 1.4 |
| A → U: MUT3-1 (C935N) | |||||
| MUT3-1 | A6U | CCAGAAUUGUAUUUAUUCG | 0.50 ± 0.099 | 4 | 1 |
| MUT3-1 | WT RNA | CCAGAAUUGUAUAUAUUCG | 2.8 ± 1.3 | 3 | 5.6 |
| G → U, A → U: MUT7-2/3-1 (C935N/S1079N/E1083Q) | |||||
| MUT7-2/3-1 | ARE | UUAUUUAUUUAUUUAUU | 0.60 ± 0.26 | 6 | 1 |
| MUT7-2/3-1 | WT RNA | CCAGAAUUGUAUAUAUUCG | 280 ± 49 | 4 | 467 |
*RNA bases recognized by the eight PUM repeats are underlined and the numbers above represent the PUM repeat that recognizes that base. Boldface letters note positions that are changed relative to wild-type NRE RNA.
†Krel represents either the affinity of wild-type protein for the mutant RNA relative to the affinity of wild-type protein for wild-type RNA or the affinity of mutant protein for wild-type RNA relative to the affinity of mutant protein for its cognate RNA.