The number of subunits comprising the channel formed by the T domain of diphtheria toxin

M. Gordon, A. Finkelstein

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

In the presence of a low pH environment, the channel-forming T domain of diphtheria toxin undergoes a conformational change that allows for both its own insertion into planar lipid bilayers and the translocation of the toxin's catalytic domain across them. Given that the T domain contributes only three transmembrane segments, and the channel is permeable to ions as large as glucosamine+ and NAD-, it would appear that the channel must be a multimer. Yet, there is substantial circumstantial evidence that the channel may be formed from a single subunit. To test the hypothesis that the channel formed by the T domain of diphtheria toxin is monomeric, we made mixtures of two T domain constructs whose voltage-gating characteristics differ, and then observed the gating behavior of the mixture's single channels in planar lipid bilayers. One of these constructs contained an NH2-terminal hexahistidine (H6) tag that blocks the channel at negative voltages; the other contained a COOH-terminal H6 tag that blocks the channel at positive voltages. If the channel is constructed from multiple T domain subunits, one expects to see a population of single channels from this mixture that are blocked at both positive and negative voltages. The observed single channels were blocked at either negative or positive voltages, but never both. Therefore, we conclude that the T domain channel is monomeric.

Original languageEnglish (US)
Pages (from-to)471-480
Number of pages10
JournalJournal of General Physiology
Volume118
Issue number5
DOIs
StatePublished - 2001

Fingerprint

Diphtheria Toxin
His-His-His-His-His-His
Lipid Bilayers
Glucosamine
NAD
Catalytic Domain
Ions
Population

Keywords

  • Histidine tags
  • Monomer
  • Planar lipid bilayers
  • Single channels
  • Voltage gating

ASJC Scopus subject areas

  • Physiology

Cite this

The number of subunits comprising the channel formed by the T domain of diphtheria toxin. / Gordon, M.; Finkelstein, A.

In: Journal of General Physiology, Vol. 118, No. 5, 2001, p. 471-480.

Research output: Contribution to journalArticle

@article{0814e6e441fb4bedac68fa3d3bd6728d,
title = "The number of subunits comprising the channel formed by the T domain of diphtheria toxin",
abstract = "In the presence of a low pH environment, the channel-forming T domain of diphtheria toxin undergoes a conformational change that allows for both its own insertion into planar lipid bilayers and the translocation of the toxin's catalytic domain across them. Given that the T domain contributes only three transmembrane segments, and the channel is permeable to ions as large as glucosamine+ and NAD-, it would appear that the channel must be a multimer. Yet, there is substantial circumstantial evidence that the channel may be formed from a single subunit. To test the hypothesis that the channel formed by the T domain of diphtheria toxin is monomeric, we made mixtures of two T domain constructs whose voltage-gating characteristics differ, and then observed the gating behavior of the mixture's single channels in planar lipid bilayers. One of these constructs contained an NH2-terminal hexahistidine (H6) tag that blocks the channel at negative voltages; the other contained a COOH-terminal H6 tag that blocks the channel at positive voltages. If the channel is constructed from multiple T domain subunits, one expects to see a population of single channels from this mixture that are blocked at both positive and negative voltages. The observed single channels were blocked at either negative or positive voltages, but never both. Therefore, we conclude that the T domain channel is monomeric.",
keywords = "Histidine tags, Monomer, Planar lipid bilayers, Single channels, Voltage gating",
author = "M. Gordon and A. Finkelstein",
year = "2001",
doi = "10.1085/jgp.118.5.471",
language = "English (US)",
volume = "118",
pages = "471--480",
journal = "Journal of General Physiology",
issn = "0022-1295",
publisher = "Rockefeller University Press",
number = "5",

}

TY - JOUR

T1 - The number of subunits comprising the channel formed by the T domain of diphtheria toxin

AU - Gordon, M.

AU - Finkelstein, A.

PY - 2001

Y1 - 2001

N2 - In the presence of a low pH environment, the channel-forming T domain of diphtheria toxin undergoes a conformational change that allows for both its own insertion into planar lipid bilayers and the translocation of the toxin's catalytic domain across them. Given that the T domain contributes only three transmembrane segments, and the channel is permeable to ions as large as glucosamine+ and NAD-, it would appear that the channel must be a multimer. Yet, there is substantial circumstantial evidence that the channel may be formed from a single subunit. To test the hypothesis that the channel formed by the T domain of diphtheria toxin is monomeric, we made mixtures of two T domain constructs whose voltage-gating characteristics differ, and then observed the gating behavior of the mixture's single channels in planar lipid bilayers. One of these constructs contained an NH2-terminal hexahistidine (H6) tag that blocks the channel at negative voltages; the other contained a COOH-terminal H6 tag that blocks the channel at positive voltages. If the channel is constructed from multiple T domain subunits, one expects to see a population of single channels from this mixture that are blocked at both positive and negative voltages. The observed single channels were blocked at either negative or positive voltages, but never both. Therefore, we conclude that the T domain channel is monomeric.

AB - In the presence of a low pH environment, the channel-forming T domain of diphtheria toxin undergoes a conformational change that allows for both its own insertion into planar lipid bilayers and the translocation of the toxin's catalytic domain across them. Given that the T domain contributes only three transmembrane segments, and the channel is permeable to ions as large as glucosamine+ and NAD-, it would appear that the channel must be a multimer. Yet, there is substantial circumstantial evidence that the channel may be formed from a single subunit. To test the hypothesis that the channel formed by the T domain of diphtheria toxin is monomeric, we made mixtures of two T domain constructs whose voltage-gating characteristics differ, and then observed the gating behavior of the mixture's single channels in planar lipid bilayers. One of these constructs contained an NH2-terminal hexahistidine (H6) tag that blocks the channel at negative voltages; the other contained a COOH-terminal H6 tag that blocks the channel at positive voltages. If the channel is constructed from multiple T domain subunits, one expects to see a population of single channels from this mixture that are blocked at both positive and negative voltages. The observed single channels were blocked at either negative or positive voltages, but never both. Therefore, we conclude that the T domain channel is monomeric.

KW - Histidine tags

KW - Monomer

KW - Planar lipid bilayers

KW - Single channels

KW - Voltage gating

UR - http://www.scopus.com/inward/record.url?scp=0035169411&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0035169411&partnerID=8YFLogxK

U2 - 10.1085/jgp.118.5.471

DO - 10.1085/jgp.118.5.471

M3 - Article

C2 - 11696606

AN - SCOPUS:0035169411

VL - 118

SP - 471

EP - 480

JO - Journal of General Physiology

JF - Journal of General Physiology

SN - 0022-1295

IS - 5

ER -