Table 1.
Bactericidal activity of epitope insertion mutants in colicin Ia
| Mutant | Amino acid residue preceding insert | Inserted sequence | Protein yield, mg/liter | Specific activity, killing units/mg |
|---|---|---|---|---|
| Wild type | NA | None | 30 | 1 × 107 |
| L474 → C | NA | None | 7 | 1 × 107 |
| HA-I | 511 | YPYDVPDYA | 14 | 1 × 106 |
| HA-II | 508 | CYPYDVPDYARI | 5 | 1 × 105 |
| HA-III | 508 | NYPYDVPDYARI | 8 | 1 × 106 |
| FLAG-IV | 508 | DYKDDDDKRI | 7 | 1 × 105 |
| FLAG-V | 508 | DYKDDDDKRI | 36 | 1 × 106 |
Epitope insertion mutants were created as described in Materials and Methods. The single-letter codes for the inserted sequences are indicated. The HA and FLAG epitope sequences are shown in bold. Specific activity was determined by spot-testing 10-fold dilutions on lawns of sensitive E. coli. Mutants HA-II, HA-III, FLAG-IV, and FLAG-V contained the mutations K507 → T and N509 → D. Mutants HA-II, HA-III, and FLAG-IV also contained the mutation L474 → C. Wild-type colicin Ia and the mutant L474 → C are shown for comparison.