Recent discovery of the non-canonical binding site on cholera toxin subunit B (CTB) showed that the CTB has affinity to several fucosylated molecules apart from the well-known canonical site binding GM1 ganglioside, independently initiate the intoxication process. Increased blocking of CTB binding to human and murine small intestine epithelial cells can be achieved when targeting both canonical and non-canonical binding sites simultaneously using multivalent norbornenyl glycopolymers. However, the mechanism behind increased blocking efficacy by concurrent targeting multiple binding sites has been remained unclear. We have witnessed CTB forms large aggregate with glycopolymers accompanying certain sugar composition. Here, we propose the mechanism of self-assembly process between glycopolymer and CTB. Our results from dynamic light scattering (DLS), small-angle X-ray scattering (SAXS), transmission electron microscopy (TEM) and turbidimetric assay have identified the facial directionality of CTB plays key role promoting interpolymer crosslinking. The interpolymer crosslinking induced self-assembly is exclusively present when the polymer system contains both galactose and fucose together. Direct correlation between IC50 and the size of self-assembly was observed. Our research highlights the significance of how macromolecular interaction affects pharmacodynamics. The ultimate advantage of such mechanism is that the blocking of CTB binding by multivalent polymeric inhibitors circumvents the need of exotic architectures thus brings insight into potential low-cost macromolecular therapeutics for clinical stage cholera treatment.

Facial directionality induced interpolymer crosslinking promotes hyperbranched self-assembly of cholera toxin B and glycomimetic polymer

ACS Fall 2020 Virtual Meeting

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