Self-translocation of diphtheria toxin across model membranes

J. X. Jiang, L. A. Chung, E. London

Research output: Contribution to journalArticle

20 Scopus citations

Abstract

To understand the mechanism of diphtheria toxin membrane translocation, the toxin was entrapped within lipid vesicles, and its low pH-induced translocation across the lipid bilayer was measured. Proteolysis and resistance to guanidinium chloride denaturation were used to demonstrate that the toxin molecules were entrapped. Low pH-induced movement of entrapped toxin to the outer (trans) face of the bilayer was assayed by the binding of external streptavidin to biotin-labeled entrapped toxin. Complete translocation was quantified by the amount of protein released into the external medium. Using whole toxin, it was found that the A fragment was efficiently translocated, but the B fragment was not. This was true both in the low temperature (A domain folded) and high temperature (A domain unfolded) toxin conformations previously identified [Jiang J.X., Abrams, F.S., and London, E. (1991) Biochemistry 30, 3857-3864]. Remarkably, even isolated fragment A appeared to self-translocate under some conditions. Toxin-induced translocation may partly result from formation of a nonspecific toxin-induced pore. This idea is supported by the toxin-induced release of fluorescent dextrans coentrapped within the vesicles. However, low pH-induced exposure of entrapped toxin on the outside of the membrane was conformation dependent. Exposure was greatest for the high temperature conformation. This suggests the existence of a more specific translocation process. The nature and relationship of these processes, and their relative roles in translocation in vivo are discussed.

Original languageEnglish (US)
Pages (from-to)24003-24010
Number of pages8
JournalJournal of Biological Chemistry
Volume266
Issue number35
StatePublished - 1991
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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