Design and Synthesis of High-Avidity Tetravalent Glycoclusters as Probes for Sambucus sieboldiana Agglutinin and Characterization of their Binding Properties

Abstract

We designed and synthesized three tetravalent sialo-glycoclusters that had different separations between the terminal sialic acids and the linking carboxy groups of the ethylene glycol bis(β-aminoethyl ether)-N,N,N',N'-tetraacetate scaffold to serve as ligands for the sialic acid-binding lectin Sambucus sieboldiana agglutinin (SSA). The interaction between each glycocluster and SSA was characterized by hemagglutination inhibition, quantitative precipitation, and double-diffusion assays. For the precipitation assays, the precipitin curves indicated that the ligands and SSA bound in either a 1:1 or a 1:2 ratio, i.e., stoichiometrically. The strong interactions of these sialo-glycoclusters with SSA could be ascribed to a combination of multivalency and spacer effects. We also assessed the nature of the ligand-SSA complexes by isothermal titration calorimetry and dynamic light scattering. The results of those experiments indicated that formation of intermolecular complexes occurred at less than stoichiometric ratios of ligand to SSA concentrations and that, as the concentrations of the ligands increased, larger cross-linked aggregates formed. Large aggregates that were present concurrently with visible precipitation and with a particle size centered at ~600 to 800 nm were identified by dynamic light scattering.

Role of the Carbohydrate-Binding Sites of Griffithsin in the Prevention of DC-SIGN-Mediated Capture and Transmission of HIV-1

“…This phenomenon and the importance of the capability of CBAs such as GRFT to concomitantly cross-link several glycans to exert efficient antiviral activity is also evident from our findings that GRFT and its single mutant variants show virtually similar low K D values (binding affinity) for gp120, but have strikingly different antiviral potencies (wild-type GRFT is by far superior to the mutant GRFT variants in its antiviral efficacy) ([14], and data in this study). The importance of cross-linking glycans on gp120 by CBAs has not only be recently shown for GRFT ([14] and this study), but also for other CBAs such as the prokaryotic cyanovirin [31], the plant lectins of Sambucus sieboldiana , Soybean, Wisteria floribunda and Vicia villosa [32], [33], the snail lectin of Helix pomatia [33] and the mammalian galectins [34].…”

Agglutination of Human Polyomaviruses by Using a Tetravalent Glycocluster as a Cross-Linker

“…Tetravalent asialo- and sialo-glycosides ( 3 , 4 , 5 , and 6 ) were prepared by our previously described method (see the Supporting Information ). 36 …”

“… 36 This lectin is a heterotetrameric molecule with a hydrodynamic diameter of 12.2 nm and two sialic acid-binding sites at its extremities. 36 , 40 , 41 In the present study, this ligand–lectin interaction was considered appropriate for the present ligand–virus from the viewpoint of the size of both acceptors. The hydrodynamic diameter of MCPyV-LP is 3.7 times larger than that of SSA.…”

Middle-molecular-weight Glycoclusters for the Crosslinking of Multivalent Lectins

“…This not only reduces the solubility of the ligands, but also limits the lectins that can be targeted. Therefore, we developed a new middle-molecular-weight glycocluster for Sambucus sieboldiana agglutinin (SSA), which has a wide sugar-binding site that can interact with multiple sugar residues in the sugar chain (49). SSA is a plant lectin present in the bark of S. sieboldiana, and is a heterotetrameric glycoprotein with two sugarbinding sites (50)(51)(52).…”

“…Here, we describe a useful SSA cross-linking agent that we have developed by controlling the valency and the spatial arrangement of ligand sugar chains. (12), in which the length corresponding to one repetition of LacNAc of 11 was replaced with an alkyl spacer (49), was also synthesized (Fig. 2).…”