Lipopolysaccharides (LPSs) are glycolipids that represent an essential component of the outer membrane of most Gram-negative bacteria. LPS molecules possess complex architectures, where a more conserved lipid domain (lipid A) is linked to a hypervariable long-chain O-antigen polysaccharide (OPS) via a region known as the core oligosaccharide. Assembly of LPS molecules and their insertion into the outer membrane requires a collection of enzymes and structural proteins that synthesize the component parts in the cytosol and span the cell envelope to translocate nascent molecules to the cell surface. Glycosylation of lipid A-core with OPS occurs in the periplasm, prior to translocation of LPS molecules to the cell surface. Glycosylation is not required for viability of laboratory-grown bacteria but OPSs are important virulence determinants that play pivotal roles in protecting many pathogens from immune defenses in the host. OPS chain length is an important element in these functions. Most OPSs are assembled one of two widespread mechanisms with fundamentally different strategies for chain-length control. In one approach, OPS biosynthesis occurs in the cytoplasm and a pathway-defining ABC-transporter exports the nascent glycan to the periplasm for glycoslation of lipid A-core. In a subset of these "ABC transporter-dependent" pathways, OPS synthesis and export are obligatorily coupled and their coordination dictates chain length. In the other subset, OPS chain length is established prior to export and involves the activities of a polymerase that extends the repeat-unit glycan and a terminator that caps its non-reducing end to block further extension. The polymerase and terminator can be contributed by independent proteins operating in a complex, or by a single polypeptide that incorproates both activities. In either scenario, a coiled-coil structure acts as a molecular ruler determines OPS chain length, likely by spatially separating the polymerase and terminator active sites. To ensure that the established chain lengths are then replicated in the completed LPS molecules, the ABC transporters that deliver the OPSs to the periplasm include a quality control system, where carbohydrate-binding modules linked to the ABC transporters select only properly terminated OPS for export. Together, these systems provide a platform for understanding chain-length regulation in other bacterial glycans.

Polymerases, terminators, molecular rulers and transporters – integrated quality control systems for establishing the chain length of LPS O antigens

253rd National Meeting (2017)

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