TABLE 1.
Characterization of wild-type and mutant IRP3 and IRP1 operators
| Plasmid | Sequence of operatora | Iron | β-Galactosidase activity (Miller units)b
|
DtxR bindingc to operator | |
|---|---|---|---|---|---|
| dtxR+ | DtxR− | ||||
| Consensus sequenced | TTAGGTTAGCCTAACCTAA | ||||
| pIRP3-1 | TTAGGTGAGACGCACCCAT | + | 5.2 ± 1.3 | 45.3 ± 2.3 | + |
| − | 46.7 ± 3.8 | 43.5 ± 1.7 | |||
| pIRP3 C(+7)T | TTAGGTGAGACGCACC*TAT | + | 0.2 ± 0.2 | 0.2 ± 0.1 | ++++ |
| − | 0.3 ± 0.3 | 0.4 ± 0.2 | |||
| pIRP3 C(+7)A | TTAGGTGAGACGCACC*AAT | + | 1.7 ± 0.5 | 6.2 ± 1.7 | ++ |
| − | 4.8 ± 1.2 | 7.4 ± 1.5 | |||
| pIRP3 C(+7)G | TTAGGTGAGACGCACC*GAT | + | 1.0 ± 0.1 | 3.1 ± 0.4 | ± |
| − | 2.8 ± 0.3 | 3.7 ± 0.9 | |||
| pIRP3 A(−7)C | TT*CGGTGAGACGCACCCAT | + | 56.4 ± 5.5 | 75.1 ± 6.4 | ± |
| − | 75.5 ± 4.1 | 79.3 ± 5.3 | |||
| pIRP3 A(−7)G | TT*GGGTGAGACGCACCCAT | + | 0.6 ± 0.3 | 1.7 ± 0.9 | ± |
| − | 2.5 ± 1.2 | 1.5 ± 0.5 | |||
| pIRP3 A(−7)T | TT*TGGTGAGACGCACCCAT | + | 1.0 ± 0.2 | 4.5 ± 0.9 | + |
| − | 3.6 ± 0.7 | 4.1 ± 1.1 | |||
| pIRP3 C(+7)T/A(−7)G | TT*GGGTGAGACGCACC*TAT | + | 0.1 ± 0.1 | 0.3 ± 0.2 | +++ |
| − | 0.3 ± 0.2 | 0.2 ± 0.1 | |||
| pIRP1-1 | TTAGGTTAGCCAAACCTTT | + | 1.1 ± 0.2 | 19.5 ± 1.8 | +++++ |
| − | 20.3 ± 2.1 | 22.3 ± 2.6 | |||
| pIRP1 T(+7)C | TTAGGTTAGCCAAACC*CTT | + | 12.7 ± 0.6 | 95.1 ± 3.8 | +++ |
| − | 87.0 ± 4.4 | 91.5 ± 4.2 | |||
| pQF50 | + | 0.2 ± 0.1 | 0.2 ± 0.1 | ||
| − | 0.1 ± 0.1 | 0.2 ± 0.2 | |||
a
Asterisks above operator sequences identify locations of nucleotide substitutions.
b
Average of at least three determinations for E. coli DH5α ± standard deviation.
c
Relative binding of DtxR to the operators from IRP3, IRP1, and their mutants, based an gel mobility shift data from Fig. 2 (pIRP3 was assigned an arbitrary value of +, and pIRP1 was assigned an arbitrary value of +++++).
d
19-bp consensus sequence for DtxR-specific operators (see text).