Multivalent Carbohydrate Recognition on a Glycodendrimer‐Functionalized Flow‐Through Chip

Abstract

Dendrimers were fitted out with up to eight mannose moieties by "click" chemistry. They were subsequently attached to aluminum oxide chips via a spacer that was linked to the dendrimer core; this resulted in a microarray of glycodendrimers. Binding of the glycodendrimers to the fluorescent lectins ConA and GNA was observable in real time. In a single experiment it was possible to observe the multivalency enhancement or cluster effect in the binding event. This effect was small for ConA, in agreement with its widely spaced binding sites, whereas it was large for GNA, with its twelve much more closely spaced binding sites. The dendrimer-fitted chip represents a valuable screening tool for multivalency effects. Furthermore kinetic and thermodynamic data on binding events can be deduced. Inhibition experiments are also possible with the system as was shown for ConA with alpha-methyl mannose as the inhibitor.

Rapid Screening of Lectins for Multivalency Effects with a Glycodendrimer Microarray

“…The picture is very similar to that obtained with our previous microarray that only contained mannosides. [13] Furthermore, the relative intensities of the different valencies is the same with FITC-labeled ConA of different commercial sources, further supporting the reproducibility of the experiments.…”

“…They were subsequently immobilized onto the maleimide-functionalized aluminum oxide microarray slides surface by using piezoelectric spotting of 330 pL per spot at pH 9. The sugar concentration of the spotting was varied from 0.5 to 5 mm, and as before, [13] a correction was made for the valency.…”

“…Compounds 1-4 a, [13] which are alkynes ranging in valency from 1 to 8, were conjugated to the azide-containing sugars 5-9 by CuAAC click chemistry according to our previously reported conditions. [16] The obtained products were subsequently treated with NaOMe to liberate the protected hydroxyl groups then treated with trifluoroacetic acid (TFA) to deprotect the amine function in the core of the structures for coupling to the chip surface.…”

“…[15] In our case, the long pores of the chip material also seemed to further enhance the binding phenomena due to the increased rebinding possibilities. [13] Multivalency effects for concanavalin A and the GNA lectin from snowdrop with the porous aluminum oxide displayed mannosides were small and large, respectively. This correlated with their potential to allow chelation, as estimated by the inter-binding-site distance, which is only possible for GNA.…”

“…Microarrays are increasingly being explored in the study of protein-carbohydrate interactions. [11,12] In order to specifically study multivalency effects, glycodendrimers were used and attached to porous aluminum oxide flowthrough microarray chips [13] that had been used successfully for recognition studies of the other major classes of biomolecules. [14] Although this was only done for mannose-containing dendrimers, it was shown that in a single experiment, an evaluation could be performed that provided information about the magnitude of multivalency effects for dendrimers ranging in valencies of up to eight.…”