Characterization of the epitope for 4C4.1 mAb on alpha-latrotoxin using phage display-peptide libraries: Prevention of toxin-dependent Ca-45(2+) uptake in nonneuronal human embryonic cells transiently expressing latrophilin

alpha -latrotoxin, a protein toxin present in the venom of black widow spid er, interacts with membrane receptors of neurons and other secretary cells to stimulate exocytosis. Two types of receptors have been identified and cl oned. Our attention has been focused on the calcium independent receptor, a G-protein coupled receptor, named latrophilin to see whether alpha -latrot oxin interaction was capable to produce an ionotropic effect, in alternativ e to the metabotropic hypothesis. Expression of latrophilin receptor is suf ficient for the alpha -latrotoxin effect to become manifest. By inducing th e transient expression of latrophilin receptor in non-neuronal human embryo nic cells, we made them susceptible to toxin action as demonstrated by the increase in Ca-45(2+) accumulation detected after toxin treatment. Since th e presence of a monoclonal antibody against alpha -latrotoxin (4C4.1 mAb) w as able to obliterate toxin-dependent effects,, we further investigated the nature of toxin-antibody interaction by characterization of the binding ep itope using phage display-peptide libraries. A conformational epitope was r ecognized and partially localized on a region of the peptide toxin whereby a tetrameric structure is formed and inserted into the membrane of target c ells where it functions as a pore. (C) 2000 Societe francaise de biochimie et biologie moleculaire / Editions scientifiques et medicales Elsevier SAS.