ABSTRACT
Protein toxins produced by bacteria are the cause of many life-threatening diarrheal diseases. Many of these toxins, including cholera toxin (CT), enter the cell by first binding to glycolipids in the cell membrane. Inhibiting these multivalent protein/carbohydrate interactions would prevent the toxin from entering cells and causing diarrhea. Here we demonstrate that the site-specific modification of a protein scaffold, which is perfectly matched in both size and valency to the target toxin, provides a convenient route to an effective multivalent inhibitor. The resulting pentavalent neoglycoprotein displays an inhibition potency (IC50) of 104â pM for the CT B-subunit (CTB), which is the most potent pentavalent inhibitor for this target reported thus far. Complexation of the inhibitor and CTB resulted in a protein heterodimer. This inhibition strategy can potentially be applied to many multivalent receptors and also opens up new possibilities for protein assembly strategies.
Subject(s)
Bacteria/metabolism , Cholera Toxin/chemistry , Binding Sites , Carbohydrates , Glycoproteins , Models, Molecular , ProteinsABSTRACT
Eight symmetric and pentavalent corannulene derivatives were functionalized with galactose and the ganglioside GM1-oligosaccharide (GM1os) via copper-catalyzed alkyne-azide cycloaddition (CuAAC) reactions. The compounds were evaluated for their ability to inhibit the binding of the pentavalent cholera toxin to its natural ligand, ganglioside GM1. In this assay, all ganglioside GM1os-sym-corannulenes proved to be highly potent nanomolar inhibitors of cholera toxin.
Subject(s)
Antitoxins/chemistry , Cholera Toxin/antagonists & inhibitors , G(M1) Ganglioside/analogs & derivatives , Galactose/chemistry , Polycyclic Aromatic Hydrocarbons/chemistry , Antitoxins/pharmacology , Cholera Toxin/metabolism , G(M1) Ganglioside/metabolism , G(M1) Ganglioside/pharmacology , Galactose/pharmacology , Polycyclic Aromatic Hydrocarbons/pharmacologyABSTRACT
Cholera toxin (CT), the causative agent of cholera, displays a pentavalent binding domain that targets the oligosaccharide of ganglioside GM1 (GM1os) on the periphery of human abdominal epithelial cells. Here, we report the first GM1os-based CT inhibitor that matches the valency of the CT binding domain (CTB). This pentavalent inhibitor contains five GM1os moieties linked to a calix[5]arene scaffold. When evaluated by an inhibition assay, it achieved a picomolar inhibition potency (IC50 = 450 pM) for CTB. This represents a significant multivalency effect, with a relative inhibitory potency of 100,000 compared to a monovalent GM1os derivative, making GM1os-calix[5]arene one of the most potent known CTB inhibitors.
Subject(s)
Antitoxins/chemistry , Antitoxins/pharmacology , Calixarenes/chemistry , Calixarenes/pharmacology , Cholera Toxin/antagonists & inhibitors , G(M1) Ganglioside/chemistry , G(M1) Ganglioside/pharmacology , Cholera/drug therapy , Cholera/microbiology , Cholera Toxin/metabolism , Humans , Models, Molecular , Vibrio cholerae/drug effects , Vibrio cholerae/enzymologyABSTRACT
Self-assembly is one of nature's mechanisms by which higher order structures are obtained. Two of the main driving forces for self-assembly, hydrophobic interactions and hydrogen bonding, are both present within amphiphilic peptides. Here, it is demonstrated how the intricately interconnected folding and assembly behavior of an N-terminally acylated peptide, with the sequence GANPNAAG, has been tuned by varying its hydrophobic tail and thermal history. The change in interplay between hydrophobic forces and peptide folding allowed the occurrence of different types of aggregation, from soluble peptides with a random coil conformation to aggregated peptides arranged in a beta-sheet assembly, which form helically twisted bilayer ribbons.