TOWARDS A RECOMBINANT VACCINE AGAINST DIPHTHERIA-TOXIN

Two recombinant fragments of diphtheria toxin (DT) were fused to an en gineered tandem repeat of the immunoglobulin (Ig) binding domain of pr otein A, called ZZ. These fragments are (i) the receptor binding domai n (DTR), which comprises amino acids 382 to 535 of DT, and (ii) a line ar peptide (DT168-220) which comprises residues 168 to 220 of the loop between fragment A and fragment B of DT. The fusion proteins were pro duced in Escherichia coli and purified by affinity chromatography. In vitro experiments showed that the DTR domain is responsible for the ca pacity of ZZ-DTR to bind to Vero cells and is capable of inhibiting th e cytotoxicity of DT for these cells. These findings suggest that DTR binds to the cell surface receptors of DT and hence adopts a conformat ion that is similar to that of the receptor binding domain of DT. We c ompared the capacities of ZZ-DTR, ZZ-DT168-220 and a chemically detoxi fied form of DT currently used for vaccination to elicit antibodies in rabbits. The toroid was more immunogenic than ZZ-DT168-220, which in turn was more immunogenic than ZZ-DTR. However, ZZ-DT168-220 antiserum was poorly efficient at neutralizing DT cytotoxicity on Vero cells, w hereas ZZ-DTR antiserum was only 15-fold less potent than anti-DT anti sera.