a, Non-reducing SDS–PAGE showing the purified LptD mutant proteins containing different Cys-to-Ser mutations. CCCC indicates that amino acids present at positions 31, 173, 724 and 725 in LptD are cysteine residues. LptD with mutation to Ser at different positions in the LptD sequence are co-expressed with LptE–His. b, Reducing SDS–PAGE showing LptD mutant proteins containing different Cys-to-Ser mutations. Samples were reduced by addition of 10 mM β-mercaptoethanol in the SDS–PAGE loading dye. All samples were boiled for 10 min at 95 °C. The degradation bands that appear below wild-type LptD were confirmed to be LptD fragments by mass spectrometry. All data shown are representative of at least three independent experiments.