Preparation and biological evaluation of self-assembled cubic phases for the polyvalent inhibition of cholera toxin

Abstract

The inverse cubic phase derived from the self-assembly of surfactants in water offers a unique three-dimensional platform for protein binding. Colloidally stable, sub-micron dispersions of the inverse bicontinuous cubic phase (cubosomes) impart an unusually large interfacial area for presentation of small molecules to selectively bind proteins of interest. Cubosomes of the phytantriol/water system were prepared and the receptor for cholera toxin (CT), monosialoganglioside G_M1 (G_M1), was integrated within the cubic phase. Our results show that G_M1-functionalised cubosomes display a strong inhibitory response against CT with a high specificity for the toxin. Surface plasmon resonance (SPR) studies demonstrate that CT and cholera toxin B subunit (CT_B) both specifically bind and form a very stable complex with G_M1–phytantriol cubosomes, demonstrated by an absence of binding to control proteins (mouse IgG, lysozyme and ricin). The inhibitory activity of the G_M1–phytantriol cubosomes against CT was evaluated by a modified enzyme linked immunosorbent assay (ELISA). Using this method we have determined a nanomolar inhibitory activity (e.g., IC₅₀ = 2.31 nM against 10 ng ml⁻¹ CT) for these particles and a dissociation constant of the G_M1–CT complex (K_D) of 1.75 nM, highlighting the remarkable inhibitory activity of the self-assembled cubic phase systems.