A linear polymer of glucose molecules, is the most abundant biopolymer on earth. It forms the major component of plant cell walls where it confers stability to the cell to allow growth based on internal pressure. Bacteria, mainly Gram-negative species, also form cellulose when transitioning from growth in solution to a sessile form (biofilms). We seek to determine the diverse mechanisms of cellulose biosyntheses in bacteria and plants. While most bacteria produce so-called amorphous cellulose, plants organize the cellulose polymers into cable-like structures, termed cellulose microfibrils, that are wrapped around the cell to form the load-bearing wall component.
Left: Cellulose is synthesized by provessive membrane-integrated glycosyltransferases that synthesize and secrete the polysaccharide. Right: Check out our latest publication in Science. Plant cellulose synthases assemble into trimeric complexes to produce cellulose protofibrils. Protofibrils likely assemble into microfibrils that form the load-bearing component of plant cell walls.
Hyaluronan (HA) is a major component of the extracellular matrix in vertebrates and affects numerous physiological processes, such as cell adhesion and migration as well as cell differentiation and proliferation. HA is a linear polysaccharide consisting of alternating glucuronic acid and N-acetylglucosamine residues and can reach several microns in length. In eukaryotes as well as in some Gram-positive bacteria, the polymer is synthesized by a membrane-embedded hyaluronan synthase (HAS) that also transports the polymer to the cell surface. We seek to determine the mechanism by which HAS accomplishes these tasks and controls HA polymer length.
HAS is a bifunctional glycosyltransferase that polymerizes UDP-activated glucuronic acid and N-acetylglucosamine linked via beta-1,4 and beta-1,3 linkages, respectively. Similar to cellulose synthase, the enzyme secretes the synthesized high molecular weight polymer across the plasma membrane during biosynthesis. Left: Expression of streptococcal HAS in E. coli leads to accumulation of HA on the cell surface. Right: Bacterial and viral HAS synthesize the same HA polymer, yet use a fundamentally different elongation mechanism. The bacterial enzyme operates as an obligate dimer in which two subunits form a single active site. A monomeric viral enzyme is sufficient for HA biosynthesis.
O antigens are complex carbohydrates that represent the variable region of lipopolysaccharides in the outer membrane of Gram-negative bacteria. The polymers form extended barriers around the cell, thereby reducing the efficacy of the host's innate immune response. LPS molecules are assembled in the periplasm by ligating O antigens to the lipid-A core. How the O antigen reaches the periplasm and is transferred to the lipid-A core are important unresolved questions. We are interested in studying the so-called ABC transporter-dependent pathway in which high molecular weight O antigens are transported across the inner membrane by a channel-forming ABC transporter, called WzmWzt.
ABC transporter-dependent O antigen biosynthesis. Left: O antigen polymers are assembled on undecaprenyl-phosphate lipids in a multi-step process. In some cases, the completed polysaccharide is chemically modified at its non-reducing end (black star). This modification is recognized by the associated WzmWzt ABC transporter. WzmWzt reorients the lipid-linked polymer in the inner bacterial membrane after which the O antigen chain is transferred to the lipid-A core. Right: Structure of WzmWzt in an ATP bound conformation.The transporter forms a continuous transmembrane channel (gold surface) with lateral exits towards the periplasmic lipid leaflet.
Last updated: January 2021