Abstract
Bacterial glycoproteins are associated with physiological and pathogenic functions of bacteria. It remains unclear whether bacterial glycoproteins can bind to specific classes of lectins expressed on host cells. Tannerella forsythia is a gram-negative oral anaerobe that contributes to the development of periodontitis. In this study, we aimed to find lectin-binding glycoproteins in T. forsythia. We performed affinity chromatography of wheat germ agglutinin, which binds to N-acetylglucosamine (GlcNAc) and sialic acid (Sia), and identified OmpA-like protein as the glycoprotein that has the highest affinity. Mass spectrometry revealed that OmpA-like protein contains O-type N-acetylhexosamine and hexose. Fluorometry quantitatively showed that OmpA-like protein contains Sia. OmpA-like protein was found to bind to lectins including E-selectin, P-selectin, L-selectin, Siglec-5, Siglec-9, Siglec-10, and DC-SIGN. The binding of OmpA-like protein to these lectins, except for the Siglecs, depends on the presence of calcium. N-acetylneuraminic acid (NeuAc), which is the most abundant Sia, inhibited the binding of OmpA-like protein to all of these lectins, whereas GlcNAc and mannose only inhibited the binding to DC-SIGN. We further found that T. forsythia adhered to human oral epithelial cells, which express E-selectin and P-selectin, and that this adhesion was inhibited by addition of NeuAc. Moreover, adhesion of an OmpA-like protein-deficient T. forsythia strain to the cells was reduced compared to that of the wild-type strain. Our findings indicate that OmpA-like protein of T. forsythia contains O-linked sugar chains that can mediate interactions with specific lectins. This interaction is suggested to facilitate adhesion of T. forsythia to the surface of host cells.
Highlights
Protein glycosylation generally occurs through attachment of glycan structures to proteins via asparagine residues (N-linked pathway) or serine/threonine residues (O-linked pathway)
We provide the first evidence that OmpA-like protein is a novel O-linked glycoprotein isolated by wheat germ agglutinin (WGA) affinity chromatography in T. forsythia
We further found that OmpA-like protein binds to lectins expressed on the surface of host cells, including E-selectin, P-selectin, L-selectin, sialic acid-binding immunoglobulin-type lectin (Siglec)-5, Siglec-9, Siglec-10, and dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN)
Summary
Protein glycosylation generally occurs through attachment of glycan structures to proteins via asparagine residues (N-linked pathway) or serine/threonine residues (O-linked pathway). O-Linked OmpA-Like Glycoprotein in T. forsythia post-translational modification has been recently found in bacterial species [1]. The gastrointestinal pathogen Campylobacter jejuni possesses an N-linked glycosylation pathway [2], whereas Neisseria species [3] and the gut symbiont Bacteroides fragilis [4] possesses O-linked glycosylation pathways. Protein glycosylation in bacteria is thought to occur at conserved glycosylation sequences: (D/E)X1NX2(S/T) (X1 and X2 can be any amino acids except for Pro) for N-linked glycosylation [2] and D(S/T)(A/I/L/V/M/T) for O-linked glycosylation [4, 5]. Bacterial glycoproteins are involved in many physiological functions of bacteria [1]. Several studies have shown that glycoproteins play roles in aspects of pathogenicity, such as antigenic variation [6], stimulation of host immunity [7, 8], and resistance against cleavage by host proteases [9]
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