TABLE 1.
MICs of copper for E. coli strain GR1 mutant derivatives in LB medium containing copper chloride
| Bacterial strain | Relevant genotype | Complementing gene in trans | MIC (mM) of CuCl2a |
|---|---|---|---|
| GR1 | ΔcueO | None | 3.25 |
| GR15 | ΔcueO ΔcusA::Cm | None | 1.5b |
| EC950 | ΔcueO ΔcusB | None | 1.5b |
| EC951 | ΔcueO ΔcusC | None | 1.75b |
| ECA067 | ΔcueO ΔcusC ΔtolC::Kan | None | 1.75b |
| EC933 | ΔcueO ΔcusF | None | 2.25 |
| cusA mutant strains | |||
| GR15(pECD738) | ΔcueO ΔcusA::Cm | cusA | 2.75bc |
| GR15(pECD739) | ΔcueO ΔcusA::Cm | cusA (M573I) | 1.5bc |
| GR15(pECD740) | ΔcueO ΔcusA::Cm | cusA (M623I) | 1.25bc |
| GR15(pECD741) | ΔcueO ΔcusA::Cm | cusA (M640I) | 2.75bc |
| GR15(pECD742) | ΔcueO ΔcusA::Cm | cusA (M672I) | 1.25bc |
| GR15(pECD743) | ΔcueO ΔcusA::Cm | cusA (M812I) | 2.5bc |
| GR15(pECD744) | ΔcueO ΔcusA::Cm | cusA (M738I) | 2.75bc |
| GR15(pECD745) | ΔcueO ΔcusA::Cm | cusA (M755I) | 2.5bc |
| GR15(pECD746) | ΔcueO ΔcusA::Cm | cusA (M792I) | 2.75bc |
| GR15(pECD747) | ΔcueO ΔcusA::Cm | cusA (M833I) | 2.5bc |
| GR15(pECD748) | ΔcueO ΔcusA::Cm | cusA (M875I-M878I-M881I) | 2.5bc |
| GR15(pECD768) | ΔcueO ΔcusA::Cm | cusA (D405N) | 1.5c |
| GR15(pECD769) | ΔcueO ΔcusA::Cm | cusA (E412Q) | 1.5c |
| GR15(pECD770) | ΔcueO ΔcusA::Cm | cusA (E412D) | 2.5c |
| GR15(pECD767) | ΔcueO ΔcusA::Cm | cusA (A399D) | 1.75c |
| cusF mutant strains | |||
| EC933(pECD735) | ΔcueO ΔcusF | cusF | 2.75c |
| EC933(pECD736) | ΔcueO ΔcusF | cusA (M69I-M71I) | 2.25c |
The MIC is defined as the lowest concentration at which no growth was observed. Strains were grown overnight and diluted 1:500 into fresh LB medium. After 2 h of growth at 37°C they were plated onto LB agar containing different concentrations (0.5 to 4.0 mM in steps of 0.25 mM) of CuCl2. The cusCFBA operon is fully expressed at copper concentrations above 100 μM (17). Growth was monitored after 16 h at 37°C. All experiments were done at least three times with identical results.
Colonies were small and mucoid at concentrations well below the MIC.
LB agar plates for complementation experiments contained anhydrotetracyline (200 μg/liter) to induce expression of complementing genes.