Helicobacter pylori adherence in the human gastric mucosa involves specific bacterial adhesins and cognate host receptors. Here, we identify sialyl-dimeric-Lewis x glycosphingolipid as a receptor for H. pylori and show that H. pylori infection induced formation of sialyl-Lewis x antigens in gastric epithelium in humans and in a Rhesus monkey. The corresponding sialic acid-binding adhesin (SabA) was isolated with the "retagging" method, and the underlying sabA gene (JHP662/HP0725) was identified. The ability of many H. pylori strains to adhere to sialylated glycoconjugates expressed during chronic inflammation might thus contribute to virulence and the extraordinary chronicity of H. pylori infection.

Adenovirus type 37 (Ad37) is a leading cause of epidemic keratoconjunctivitis (EKC), a severe and highly contagious ocular disease. Whereas most other adenoviruses infect cells by engaging CD46 or the coxsackie and adenovirus receptor (CAR), Ad37 binds previously unknown sialic acid-containing cell surface molecules. By glycan array screening, we show here that the receptor-recognizing knob domain of the Ad37 fiber protein specifically binds a branched hexasaccharide that is present in the GD1a ganglioside and that features two terminal sialic acids. Soluble GD1a glycan and GD1a-binding antibodies efficiently prevented Ad37 virions from binding and infecting corneal cells. Unexpectedly, the receptor is constituted by one or more glycoproteins containing the GD1a glycan motif rather than the ganglioside itself, as shown by binding, infection and flow cytometry experiments. Molecular modeling, nuclear magnetic resonance and X-ray crystallography reveal that the two terminal sialic acids dock into two of three previously established sialic acid-binding sites in the trimeric Ad37 knob. Surface plasmon resonance analysis shows that the knob-GD1a glycan interaction has high affinity. Our findings therefore form a basis for the design and development of sialic acid-containing antiviral drugs for topical treatment of EKC.

A semi-automated routine for continuous rotation electron diffraction has been developed, enabling high-throughput data collection. Serial electron crystallography combined with a deep convolutional network are used to screen for suitable crystals.

Key Points• A new histo-blood group system was discovered, based on the identification of Forssman glycolipid antigen on human red blood cells.• A newly described polymorphism in the GBGT1 gene activates the encoded enzyme to synthesize Forssman antigen. IntroductionCarbohydrate histo-blood group antigens, first recognized on red blood cells (RBCs) in 1900, 1 have been suggested to be part of our innate immune response. 2 Major carbohydrate histo-blood groups in man include the ABO, P1PK, H, Lewis, I, and GLOB systems in which glycoproteins and glycolipids carry immunodominant terminal sugars, 3 defining polymorphic antigens. Other mammals also express carbohydrate histo-blood groups, such as ABO, 4 fucoseless B antigen (Galili), 5 and Forssman (Fs) 6,7 but their expression on RBCs varies among species. Although the biologic function of polymorphic carbohydrates on RBCs is unresolved, these antigens can be used as receptors by pathogens [8][9][10][11] and their expression in tissues and bodily secretions are thus believed to reflect microbial selection. 8 In response to blood-group-mimicking glycans on bacterial surfaces, naturally occurring antibodies with the capacity to neutralize various microorganisms are formed. However, these antibodies also constitute substantial transfusion and transplantation barriers. 3,12 In 1987, 3 families with a supposed ABO subgroup named A pae were reported. 13 Although Helix pomatia lectin reacted strongly and polyclonal anti-A weakly with RBCs from some family members, monoclonal (MAb) anti-A reagents were later shown to be nonreactive, thus presenting an apparent paradox. The biochemical and genetic background of this enigmatic phenotype has remained unknown, as has its biologic consequences. We hypothesized that an explanation may be found by studying the glycolipids of this phenotype. 14 Here we report the identification of Fs glycolipids, normally found only on RBCs of selected nonprimate mammals, are strongly expressed on human A pae RBCs. In nonprimates, Fs antigen is synthesized by Fs synthase (globoside 3-␣-N-acetyl-D-galactosaminyltransferase, EC2.4.1.88), 7 an enzyme homologous to the ABO transferase. We also reveal a genetic polymorphism in the human Fs gene (GBGT1) that alters the enzymatically inactive human protein 15 Methods SamplesFive and 3 RBC units were collected from each of 2 unrelated A pae individuals (A pae #1 and A pae #2, respectively) from 2 of the originally reported families. 13 GlycolipidsGlycolipid preparation. Lysed blood units were thawed and total neutral glycolipids with Ͻ 20 sugar residues were isolated (see supplemental Methods, where control glycolipid preparations are also described; available on the Blood Web site; see the Supplemental Materials link at the top of the online article).Open-column chromatography. Total glycolipids (ϳ 110 mg) from each of A pae #1 and #2 were fractionated by silica chromatography column (5g silica/100 mg lipid; Silica 60, Merck) in a system of chloroform (C) methanol (M) solvent mixes (supplemental Metho...

Binding studies of various glycolipids, mainly belonging to the gangio series, to the toxins isolated from Vibrio cholerae, Eschienchia coil, and Closridium tetan have been performed, using the microtiter well assay. By using the found binding preferences in conjunction with minimum-energy conformations obtained from molecular modeling of the various ligands, binding epitopes on the natural receptor glycolipids for the toxins have been defined. The binding preferences for the cholera toxin and the heat-labile E. coil toxin are very similar, with the gangoide GM1 being the most efient lgd. The tetanus toxin binds strongly to ganglides of the Glb series, with GT1b as the most efficient igand. It is found that the binding epitope on GM1 for the cholera and heat-labile toxins to a large extent overlaps with the epitope on GQ1b for the tetanus toxin.The natural carbohydrate ligand for the toxin produced by Vibrio cholerae is the ganglioside GM1, which contains a single sialic acid residue (1-3). Since the discovery by van Heyningen that gangliosides are responsible for toxin binding (4), a wealth ofbiochemical and physicochemical studies have been carried out to define the important structural characteristics which determine the toxin's mode of action. The threedimensional structure of GM1 has been determined by NMR in conjunction with computational studies of its conformation (5-7), thereby allowing an approximation of the binding epitope on this glycolipid to be made. The recent crystal structure of the heat-labile enterotoxin (LT) from Escherichia coli (8) and its complex with lactose (9) provides a further impetus to the type of studies mentioned above due to the strong sequence homology (>80%o) with the cholera toxin (CT) (10, 11).