Expression and purification of His-tagged cytochrome bo3 from E. coli C43 strains Stock solutions/materials: Same as the materials needed for the purification from E. coli GO105 strain, with one additional solution: 1 M IPTG Protocol: 1) Inoculate a colony of transformed E. coli C43 wild-type bo3/E286C strain in 200 ml of LB|0.1 mg/mL carbenicillin broth (add 200 μL 100mg/mL car. into 200 mL); incubate overnight at 37 °C|200 RPM. 2) Prepare 8*500 mL of LB broth|0.1 mM CuSO4|0.1 mg/mL carbenicillin (add 500 μL 100 mM CuSO4 and 500 μl 100mg/mL car. into 500 mL in a 2 L baffled flask); sterilise, measure OD and inoculate them with 2 % starter culture (10 mL inoculums into 500 mL). 3) Incubate at 37°C/200 RPM until OD600 reaches 0.6; don’t forget to subtract LB’s OD; monitor OD hourly. 4) Add IPTG to a final concentration of 0.4 mM (200 µL of 1 M stock into 500 mL). 5) Continue the incubation for a further 3 h. After this step, follow the same procedures in the purification from E. coli GO105 strain.