Size and orientation of the lipid II headgroup as revealed by AFM imaging

In this study, we investigated the size and orientation of the bacterial Lipid II (L II) headgroup when the L II molecule is present in liquid-crystalline domains of DOPC in a supported DPPC bilayer. Using atomic force microscopy, we detected that L II causes the appearance of a 1.9 nm thick layer, situated over the DOPC headgroup region. With an increased scanning force, this layer can be penetrated by the AFM tip down to the level of the DOPC bilayer. Using different L II precursor molecules, we demonstrated that the detected layer consists of the headgroups of L II and that the MurNAc-pentapeptide unit of the headgroup is responsible for the measured 1.9 nm height of that layer. Monolayer experiments provided information about the in-plane dimensions of the L II headgroup. On the basis of these results and considerations of the molecular dimensions of L II headgroup constituents, we propose a model for the orientation of the L II headgroup in the membrane. In this model, the pentapeptide of the L II headgroup is rather extended and points away from the bilayer surface, which could be important for biological processes, in which L II is involved.