STRUCTURAL DIVERSITY IN A CONSERVED CHOLERA-TOXIN EPITOPE INVOLVED INGANGLIOSIDE BINDING

Cholera is a widespread disease for which there is no efficient vaccin e. A better understanding of the conformational rearrangements at the epitope might be very helpful for the development of a good vaccine. C holera toxin (CT) as well as the closely related heat-labile toxin fro m Escherichia coli (LT) are composed of two subunits, A and B, which f orm an oligomeric assembly AB(5). Residues 50-64 on the surface of the B subunits comprise a conserved loop (CTP3), which is involved in sac charide binding to the receptor on epithelial cells. This loop exhibit s remarkable conformational plasticity induced by environmental constr aints. The crystal structure of this loop is compared in the free and receptor-bound toxins as well as in the crystal and solution structure s of a complex with TE33, a monoclonal antibody elicited against CTP3. In the toxins this loop forms an irregular structure connecting a bet a-strand to the central alpha-helix. Ser 55 and Gln 56 exhibit conside rable conformational variability in the five subunits of the unligande d toxins. Saccharide binding induces a change primarily in Ser 55 and Gln 56 to a conformation identical in all five copies. Thus, saccharid e binding confers rigidity upon the loop. The conformation of CTP3 in complex with TE33 is quite different. The amino-terminal part of CTP3 forms a beta-turn that fits snugly into a deep binding pocket on TE33, in both the crystal and NMR-derived solution structure. Only 8 and 12 residues out of 15 are seen in the NMR and crystal structures, respec tively. Despite these conformational differences TE33 is cross-reactiv e with intact CT, albeit with a thousandfold decrease in affinity. Thi s suggests a different interaction of TE33 with intact CT.