A novel glycan modifies the flagellar filament proteins of the oral bacterium Treponema denticola

Abstract

While protein glycosylation has been reported in several spirochetes including the syphilis bacterium Treponema pallidum and Lyme disease pathogen Borrelia burgdorferi, the pertinent glycan structures and their roles remain uncharacterized. Herein, we report a novel glycan with an unusual chemical composition and structure in the oral spirochete Treponema denticola, a keystone pathogen of periodontitis. The identified glycan of mass 450.2 Da is composed of a monoacetylated nonulosonic acid (Non) with a novel extended N7 acyl modification, a 2-methoxy-4,5,6-trihydroxy-hexanoyl residue in which the Non has a pseudaminic acid configuration (L-glycero-L-manno) and is β-linked to serine or threonine residues. This novel glycan modifies the flagellin proteins (FlaBs) of T. denticola by O-linkage at multiple sites near the D1 domain, a highly conserved region of bacterial flagellins that interact with Toll-like receptor 5. Furthermore, mutagenesis studies demonstrate that the glycosylation plays an essential role in the flagellar assembly and motility of T. denticola. To our knowledge, this novel glycan and its unique modification sites have not been reported previously in any bacteria.

Flagellin Glycoproteomics of the Periodontitis Associated Pathogen Selenomonas sputigena Reveals Previously Not Described O-glycans and Rhamnose Fragment Rearrangement Occurring on the Glycopeptides

“…In the S. sputigena flagellin C9LY14, the glycosylated residues T149, S182 and T199 are very close to the N-terminal D1/β-hairpin region (Figure 2). Glycosylation of flagellin so close towards the N-terminus of the protein is unusual and has so far only been described for C. jejuni FlaA (55) and for the periplasmic flagella of the spirochete Treponema denticola, for which modification with a novel O-glycan within the conserved D1 domain has been reported very recently (56). It is tempting to speculate that glycans in the proteins' N-terminal region could potentially affect recognition of flagellin monomers by TLR5 and it will be an interesting future task to establish the role post-translational modifications of these residues play in a larger systems biology context.…”

A pleiotropic role of FlaG in regulating the cell morphogenesis and flagellar homeostasis at the cell poles of Treponema denticola

“…Treponema denticola strain ATCC 35405 (wild type) was used in this study. Cells were grown in tryptone‐yeast extract‐gelatin‐volatile fatty acids‐serum (TYGVS) medium at 37°C in an anaerobic chamber in presence of 85% nitrogen, 10% carbon dioxide, and 5% hydrogen (Kurniyati et al, ; Kurniyati, Zhang, Zhang, & Li, ). T .…”

“…denticola belongs to the phylum of spirochetes (Charon et al, ). It has 2–3 periplasmic flagella (PFs) that arise from each end of the cell and extend towards the centre where they overlap (Izard et al, ; Kurniyati et al, ). Structurally, the PFs of T .…”

“…denticola is more complicated than its counterparts in other bacteria (Charon et al, ; Li, Motaleb, Sal, Goldstein, & Charon, ). It is comprised of at least one sheath protein (FlaA) and three core proteins (FlaB1, FlaB2, and FlaB3; Kurniyati et al, ; Ruby et al, ; Seshadri et al, ). In contrast, the filaments of E .…”

“…The FlaB proteins are ~31.0–32.0 kDa in sizes and are modified with a novel glycan, which is essential for the flagellation and motility of T . denticola (Kurniyati et al, ). The FlaB proteins form a core that is sheathed by FlaA.…”