Extended Data Figure 6: Three proline residues at LptD and the proposed disulphide bond formed by the double mutation N232C and N757C, and LptD–LptE simulation stability and the separation of β1 and β26.

a, P231, P246 and P261 are located at strands β1, β2 and β3 of LptD, respectively. b, The double cysteine mutant N232C/N757C may form a disulphide bond at the lumen of the barrel, crosslinking strands β1 and β26, and preventing the lateral opening that is key to the proposed mechanism of LPS insertion. This double mutation is fatal to E. coli. c, The structural fluctuations of LptD–LptE complex over 100 ns at 323 K. Regions of high stability are coloured blue, while the more mobile domains are coloured red. Overall the barrel architecture is stable in the bilayer. d, By applying a negative constant pressure to the membrane plane it is possible to simulate the opening of the lateral gate between the strands β1 and β26. e, The LptD–LptE complex is shown from the periplasmic side at the start and 50 ns points of a simulation with a −70 bar pressure coupling. In addition to opening of the lateral gate, the extracellular region of the barrel also separates to potentially accommodate the large O-antigen and core oligosaccharide of LPS.