TABLE 2. Co-expression of FepN and Fepβ.
E. coli strain OKN3, alone or harboring plasmids, was tested for FepA-dependent functions.
| Host strain/plasmidsa | Alleleb | ColBc | ColDc |
|---|---|---|---|
| OKN3 | fepA | R | R |
| OKN3/pITS23 | fepA+ | 106 | 105 |
| OKN3/pFepN | fepN-(1–151) | R | R |
| OKN3/pFepβ | fepβ-(1–17; 151–724) | R | R |
| OKN3/pFepNpFepβ | FepNfepβ | 4 × 104 | 3 × 103 |
| OKN13 | tonBfepA | R | R |
| OKN13/pITS23 | fepA+ | R | R |
| OKN13/pFepNpFepβ | fepNfepβ | R | R |
OKN3 contains an engineered deletion of the fepA structural gene (“Experimental Procedures”), pITS23 and pFepβ are derivatives of the low copy number vector pHSG575, and pFepN is a derivative of pUC18.
The fepN allele encodes residues 1–150 of wild-type FepA, under the control of its natural promoter; fepβ (fepAΔ18–150) encodes residues 1–17 followed by a deletion of residues 18–150 and continues with residues 151–724.
Purified colicin was diluted in a microtiter plate over a 108-fold range, and 5 μ l of each dilution was transferred to an LB agar plate, previously coated with 2.5 × 108 bacterial cells in LB top agar. The plates were incubated overnight at 37 °C. The tabulated values are the reciprocal of the final dilution that showed visible clearing of the bacterial lawn.