Proton channel blockers inhibit Duox activity independent of Hv1 effects

Monica Valencia Gattas, Adam Jaffe, Juliana Barahona, Gregory E. Conner

Research output: Contribution to journalArticle

Abstract

The NADPH oxidase reaction produces protons. In the case of the NADPH oxidase, NOX2, activity depends on secretion of these protons and is inhibited by blockade of the voltage-gated proton channel (Hv1). Duox1 and Duox2 activities similarly produce intracellular protons but synthesize hydrogen peroxide directly instead of superoxide. Hv1 contributes to acid secretion in some epithelia that express Duox. To test the hypothesis that Duox activity is also sensitive to Hv1 channel blockers, Duox was assayed in the presence of either Zn2+ or 5-chloro-2-guanidinobenzimidazole (ClGBI). Both compounds inhibited Duox activity in normal human bronchial epithelial cells but with an IC50 over 10-fold higher than that reported for Hv1 (IC50 Zn2+ = 0.68 mM; IC50 ClGBI = 0.07–0.14 mM). Homogenized HEK293T cells expressing either Duox1 or Duox2 showed similar IC50 values for ClGBI suggesting these compounds inhibit the enzymes through alternate mechanisms independent of Hv1 proton secretion. Inclusion of superoxide dismutase did not restore Duox hydrogen peroxide synthesis. Addition of nigericin to eliminate any possible transmembrane pH gradients in intracellular membrane-localized Duox did not alter activity in HEK293T homogenates. Extracellular Zn2+ blocked intracellular Ca2+ increases needed for Duox activity. Together the data suggest that Duox enzyme activities in epithelia are inhibited by compounds that block Hv1 but inhibition occurs through Hv1-independent mechanisms and support the idea that Hv1 is not required for Duox activity.

Original languageEnglish (US)
Article number101346
JournalRedox Biology
Volume28
DOIs
StatePublished - Jan 2020

Fingerprint

Protons
Inhibitory Concentration 50
NADPH Oxidase
Hydrogen Peroxide
Epithelium
Nigericin
Intracellular Membranes
Proton-Motive Force
Enzyme activity
Enzymes
Superoxides
Superoxide Dismutase
Epithelial Cells
Cells
Membranes
Acids
Electric potential

Keywords

  • Bronchial epithelium
  • Duox
  • Hv1
  • Zinc

ASJC Scopus subject areas

  • Organic Chemistry
  • Clinical Biochemistry

Cite this

Proton channel blockers inhibit Duox activity independent of Hv1 effects. / Gattas, Monica Valencia; Jaffe, Adam; Barahona, Juliana; Conner, Gregory E.

In: Redox Biology, Vol. 28, 101346, 01.2020.

Research output: Contribution to journalArticle

Gattas, Monica Valencia ; Jaffe, Adam ; Barahona, Juliana ; Conner, Gregory E. / Proton channel blockers inhibit Duox activity independent of Hv1 effects. In: Redox Biology. 2020 ; Vol. 28.
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