A Single Native Ganglioside GM(1)-Binding Site Is Sufficient for Cholera Toxin To Bind to Cells and Complete the Intoxication Pathway

Jobling, Michael G.; Yang, ZhiJie; Kam, Wendy R.; Lencer, Wayne I.; Holmes, Randall K.

, upper panel, lanes 5 to 7) remained fully associated and ran near the 66-kDa marker (lane 1), whereas unboiled holotoxins with one or two CTB-G33D subunits exhibited partial dissociation into B pentamer and free CTA (Fig. 2 , upper panel, lanes 8 and 9). The high stability of the heterohexameric AB 5 holotoxins containing five, four, or three CTB-G33D polypeptides is also evident in the upper panel of Fig. 1D . As expected, increasing the number of tagged CTB polypeptides resulted in moderate decreases in observed mobilities of the pentamer bands (Fig. 2 , upper panel, compare lanes 2, 3, and 4) and the holotoxin bands (Fig. 2 , upper panel, compare lanes 5, 6, and 7), and increasing the number of CTB-G33D polypeptides resulted in slight increases in mobilities of the holotoxin bands (Fig. 2 , upper panel, compare lanes 6 to 8 and lanes 7 to 9). Upon boiling, all holotoxins dissociated and resolved into their individual polypeptide components (Fig. 2, lower panel) . All holotoxins have one CTA subunit. Holotoxins in lanes 2 and 5 are predicted to have five native CTB monomers; holotoxins in lanes 3, 6, and 8 have are predicted to have one tagged CTB and 4 untagged CTB monomers; and holotoxins in lanes 4, 7, and 9 are predicted to have two tagged and three untagged CTB monomers.