THE EFFECTS OF HELIX BREAKING MUTATIONS IN THE DIPHTHERIA-TOXIN TRANSMEMBRANE DOMAIN HELIX LAYERS OF THE FUSION TOXIN DAB(389)IL-2

The fusion protein toxin DAB(389)IL-2 is composed of the catalytic and transmembrane domains of diphtheria toxin genetically linked to human interleukin 2 (IL-2). This fusion toxin is selectively toxic for euka ryotic cells which express the high-affinity form of the IL-2 receptor and the mechanism of intoxication parallels that of native diphtheria toxin. We used site-directed mutagenesis to introduce Pro residues in to each of the three helical layers of the transmembrane domain. Altho ugh each of the mutations results in the complete loss of cytotoxic ac tivity, individual mutants were found to vary with respect to channel formation in planar lipid bilayers, binding affinity and melting tempe rature. We propose that each of the three helix layers plays a critica l role in the productive delivery of the catalytic domain to the cell cytosol.