Mutational analysis of the helical hairpin region of diphtheria toxin transmembrane domain.

摘要: Entry of the catalytic domain diphtheria toxin into cytoplasma eukaryotic cells depends on insertion T (transmembrane) endosomal membrane, a process triggered by low pH. To probe mechanism insertion, we mutated ionizable residues within helical hairpin region domain. Only three mutations caused significant effects cytotoxicity, D295K, E349K, and D352K. Each these represents substitution basic for an acidic residue at tip hairpin. Substitution Lys Glu349 or Asp352, in TH8/9 hairpin, reduced toxicity Vero > 100-fold, whereas Asp295, one 3 TH5/6/7 less marked reduction. All also altered pH-dependent formation, and/or ion conductance, channels formed artificial bilayers plasma membrane. E349K D352K did not alter pH dependence conformational changes occurring near 5. Our findings support hypothesis that inserts membrane after pH-mediated partial unfolding A positive this apparently inhibits blocks action. The ion-conducting properties selected mutants, described elsewhere, are consistent with model. status integral form remains uncertain.