Structure-function relationships in diphtheria toxin channels: I. Determining a minimal channel-forming domain

J. A. Silverman, J. A. Mindell, H. Zhan, A. Finkelstein, R. J. Collier

Research output: Contribution to journalArticle

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Abstract

Diphtheria Toxin (DT) is a 535 amino acid exotoxin, whose active form consists of two polypeptide chains linked by an interchain disulphide bond. DT's N-terminal A fragment kills cells by enzymatically inactivating their protein synthetic machinery; its C terminal B chain is required for the binding of toxin to sensitive cells and for the translocation of the A fragment into the cytosol. This B fragment, consisting of its N-terminal T domain (amino acids 191-386) and its C-terminal R domain (amino acids 387-535) is responsible for the ion-conducting channels formed by DT in lipid bilayers and cellular plasma membranes. To further delineate the channel-forming region of DT, we studied channels formed by deletion mutants of DT in lipid bilayer membranes under several pH conditions. Channels formed by mutants containing only the T domain (i.e., lacking the A fragment and/or the R domain), as well as those formed by mutants replacing the R domain with Interleukin-2 (Il-2), have single channel conductances and selectivities essentially identical to those of channels formed by wild-type DT. Furthermore, deleting the N-terminal 118 amino acids of the T domain also has minimal effect on the single channel conductance and selectivity of the mutant channels. Together, these data identify a 61 amino acid stretch of the T domain, corresponding to the region which includes α-helices TH8 and TH9 in the crystal structure of DT, as the channel-forming region of the toxin.

Original languageEnglish (US)
Pages (from-to)17-28
Number of pages12
JournalThe Journal of Membrane Biology
Volume137
Issue number1
DOIs
StatePublished - Jan 1994

Fingerprint

Diphtheria Toxin
Amino Acids
Lipid Bilayers
Exotoxins
Ion Channels
Disulfides
Cytosol
Interleukin-2
scorpion toxin I''
Cell Membrane
Peptides
Membranes
Proteins

Keywords

  • Channel-forming peptides
  • Diphtheria toxin
  • Ion channels
  • Planar lipid bilayers
  • Site-directed mutagenesis
  • T-domain

ASJC Scopus subject areas

  • Physiology
  • Cell Biology
  • Biophysics

Cite this

Structure-function relationships in diphtheria toxin channels : I. Determining a minimal channel-forming domain. / Silverman, J. A.; Mindell, J. A.; Zhan, H.; Finkelstein, A.; Collier, R. J.

In: The Journal of Membrane Biology, Vol. 137, No. 1, 01.1994, p. 17-28.

Research output: Contribution to journalArticle

Silverman, J. A. ; Mindell, J. A. ; Zhan, H. ; Finkelstein, A. ; Collier, R. J. / Structure-function relationships in diphtheria toxin channels : I. Determining a minimal channel-forming domain. In: The Journal of Membrane Biology. 1994 ; Vol. 137, No. 1. pp. 17-28.
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