Transport of lipopolysaccharide across the cell envelope: the long road of discovery

Abstract

Intracellular lipid transport is poorly understood. Genetic studies to identify lipid-transport factors are complicated by the essentiality of many lipids, whereas biochemical and cell biology approaches aiming to determine localization and mechanisms of lipid transport are often challenged by the lack of adequate technology. Here, we review the epic history of how different approaches, technological advances and ingenuity contributed to the recent discovery of a multi-protein pathway that transports lipopolysaccharide across the envelope of Gram-negative bacteria.

Figure 1. Structure of E. coli LPS

The structure of E. coli LPS with a K-12 core region is shown. The structure of LPS of several species has been solved and it is clear that it is very diverse. Moreover, one single species can further modify its typical LPS structure in response to environmental signals and defects in envelope conditions., , , Abbreviations: Kdo, 3-deoxy-D-manno-oct-2-ulosonic acid; Hep, L-glycero-D-manno-heptose; Glu, D-glucose; Gal, D-galactose; EtN, ethanolamine; P, phosphate.

Figure 2. Electron micrograph of Veillonella illustrating the ultrastructure of the Gram-negative cell envelope

Bladen and Mergenhagen published this micrograph in 1964 and they labeled the different structures as the outer membrane (OM), the solid membrane (SM), which is known nowadays as the peptidoglycan, and the plasma membrane (PM), which we refer to as the inner or cytoplasmic membrane. This figure was reproduced with the permission of the Journal of Bacteriology (pending).

Figure 3. Structure of the Gram-negative cell envelope

In Gram-negative bacteria, the cytoplasm is surrounded by the IM, a phospholipid (PL) bilayer that also contains proteins. There are two types of proteins in the IM: integral IM proteins, which span membrane through α-helical TM domains, and IM lipoproteins, which are anchored to the outer leaflet through a lipid moiety. The periplasm is the aqueous compartment bounded by the IM and the OM; it contains soluble proteins and the peptidoglycan layer. The OM is anchored to the rest of the cell via proteins that are covalently attached to the peptidoglycan. The OM is asymmetric, as it contains phospholipids in the inner leaflet and LPS in the outer leaflet. In addition, there are two types of proteins, integral OM proteins (OMPs) and lipoproteins.

Figure 4. Current models for LPS transport across the cell envelope

After being synthesized, rough LPS molecules are flipped across the IM by the ABC transporter MsbA. If the O-antigen is made, it is then ligated to the rest of the molecule by WaaL (step not shown). Then, LPS is extracted from the IM by LptC and the LptFGB ABC transporter. In the soluble-intermediate model (shown on the left), LptA serves as a soluble periplasmic chaperone that transports LPS from the IM to the LptDE assembly site. In the trans-envelope complex model (shown on the right), the Lpt factors constitute a multiprotein complex that spans the cell envelope. Because LptA crystallizes as a fiber in the presence of LPS, it is possible that it serves as a bridge between the OM and IM components. Note that the only physical interaction between Lpt factors that has been conclusively demonstrated to date is the one that exists between LptD and LptE.