FIG 3.
Complementation analysis of E. coli UQ8 biosynthesis mutants with the putative Ubi proteins from F. novicida. (A and B) The Δubi E. coli mutant strains transformed with pTrc99a (vector [vec]) or pTrc99a encompassing the ubiFn genes were grown overnight at 37°C in LB medium (A) or M9 minimal medium (B) with 0.4% (wt/vol) glucose as the sole carbon source. The expression of the UbiFn proteins was induced by the addition of IPTG to a final concentration of 100 μM. E. coli wild-type (WT) strain MG1655 transformed with the pTrc99a empty vector was used as a control. HPLC-ECD analysis of lipid extracts from 1 mg of cells was performed. The chromatograms are representative of results from three independent experiments. The peaks corresponding to OPP, DDMQ8, DMQ8, UQ8, MK8, DMK8, and the UQ10 standard are indicated. (C) Serial dilutions were spotted onto plates containing M9 minimal medium with 0.4% (wt/vol) glucose or succinate as the sole carbon source and IPTG (100 μM final concentration). The plates were incubated overnight at 37°C.