Abstract
The outer membrane (OM) of Gram-negative bacteria is a complex asymmetric bilayer containing lipids, lipopolysaccharides (LPS) and proteins. While it is a mechanical and chemical barrier, it is also the primary surface of bacterial recognition processes that involve infection by and of the bacterium. Uncovering the mechanisms of these biological functions has been hampered by the lack of suitable model systems. Here we report the step-by-step assembly of a synthetic OM model from its fundamental components. To enable the efficient formation of a supported lipid bilayer at room temperature, dimyristoyl-phosphocholine (DMPC) was used as the lipid component to which we progressively added LPS and OM proteins. The assembled system enabled us to explore the contribution of the molecular components to the topographical structure and stability of the OM. We found that LPS prefers solid-state membrane regions and forms stable vesicles in the presence of divalent cations. LPS can gradually separate from DMPC membranes to form independent vesicles, pointing at the dynamic nature of the lipid-LPS system. The addition of OM proteins from E. coli and saturating levels of LPS to DMPC liposomes resulted in a thicker and more stable bilayer the surface of which displayed a nanoscale texture formed of parallel, curved, long (>500 nm) stripes spaced apart with a 15 nm periodicity. The synthetic membrane may facilitate the investigation of binding and recognition processes on the surface of Gram-negative bacteria.
Highlights
The outer membrane (OM) of E. coli is a multi-functional asymmetrical membrane system responsible for transporting nutrients into the cytoplasm while isolating it from harmful agents in the environment
Incorporation of OM proteins extracted from E. coli further increased membrane thickness and stability, indicating that the bacterial outer membrane is mechanically reinforced by LPS and proteins
Samples were heated to 30 C which exceeds the transition temperature, at which DMPC is expected to form homogeneous uid-phase supported lipid bilayers (SLBs)
Summary
The outer membrane (OM) of E. coli is a multi-functional asymmetrical membrane system responsible for transporting nutrients into the cytoplasm while isolating it from harmful agents in the environment. Its fundamental components are lipids, lipopolysaccharides (LPS) and various types of proteins. Infection by bacteriophages such as T7,1–3 is thought to be intimately linked to the structure and properties of the OM. About 75% of the OM surface is made up of lipopolysaccharides (LPS) which are highly heterogeneous, negatively charged molecules. The remaining area is occupied by proteins.[4,5] In rough type LPS only the core region is present, whereas in smooth type variants the oligosaccharide chain is much longer, resulting in a severalnanometer-thick layer. Subgroups of rough type LPS are Re, Rd, Rc, Rb and Ra in growing oligosaccharide chain length order.[6] LPS is responsible for inducing various biological
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