Regulation of the thyroid NADPH-dependent H2O2 generator by Ca2+: Studies with phenylarsine oxide in thyroid plasma membrane

Y. Gorin, A. M. Leseney, R. Ohayon, C. Dupuy, J. Pommier, A. Virion, D. Deme

Research output: Contribution to journalArticle

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Abstract

Pig thyroid plasma membranes contain a Ca2+-dependent NADPH:O2 oxidoreductase, the thyroid NADPH-dependent H,O, generator. This provided the H,O, for the peroxidase-catalysed synthesis of thyroid hormones. The effect of the tervalent arsenical, phenylarsine oxide (PAO), on the NADPH oxidase was studied. PAO caused two directly related dose-dependent effects with similar hall-effect concentrations of PAO (3 nmol of PAO/mg of protein): (i) partial inactivation of H2O2 formation by the Ca2+-stimulated enzyme, and (ii) desensitization of the enzyme activity to Ca2+. PAO had no effect on membranes that had been Ca2+-desensitized by α-chymotrypsin treatment. The NADPH oxidase in membranes treated with excess PAO had the same V(max) with and without Ca2+. This value was half the V(max) of the native enzyme: However, the K(m) for NADPH determined with Ca2+ (18 μM, identical with that of the native enzyme) was approx. one-third of the K(m) measured without Ca2+, showing the direct action of Ca2+ on the PAO-enzyme complex. PAO had the same effects, partial inactivation and Ca2+ desensitization, on the NADPH:ferricyanide oxidoreductase activity of the NADPH oxidase, suggesting that PAO acts on the flavodehydrogenase entity of the enzyme. Both partial inactivation and Ca2+ desensitization were completely and specifically reversed by 2,3-dimercaptopropanol, partly reversed by dithiothreitol and not reversed by 2-mercaptoethanol, indicating that PAO binds to vicinal thiol groups. These results suggest that thiol groups are involved in the control of thyroid NADPH oxidase by Ca2+; PAO bound to vicinal thiols might alter the structure of the enzyme so that electron transfer occurs without Ca2+ but more slowly.

Original languageEnglish (US)
Pages (from-to)383-388
Number of pages6
JournalBiochemical Journal
Volume321
Issue number2
StatePublished - 1997
Externally publishedYes

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Cell membranes
NADP
Thyroid Gland
Cell Membrane
NADPH Oxidase
Enzymes
Sulfhydryl Compounds
Oxidoreductases
oxophenylarsine
Arsenicals
Membranes
Dimercaprol
Mercaptoethanol
Dithiothreitol
Hall effect
Chymotrypsin
Enzyme activity
Thyroid Hormones
Peroxidase
Swine

ASJC Scopus subject areas

  • Biochemistry

Cite this

Gorin, Y., Leseney, A. M., Ohayon, R., Dupuy, C., Pommier, J., Virion, A., & Deme, D. (1997). Regulation of the thyroid NADPH-dependent H2O2 generator by Ca2+: Studies with phenylarsine oxide in thyroid plasma membrane. Biochemical Journal, 321(2), 383-388.

Regulation of the thyroid NADPH-dependent H2O2 generator by Ca2+ : Studies with phenylarsine oxide in thyroid plasma membrane. / Gorin, Y.; Leseney, A. M.; Ohayon, R.; Dupuy, C.; Pommier, J.; Virion, A.; Deme, D.

In: Biochemical Journal, Vol. 321, No. 2, 1997, p. 383-388.

Research output: Contribution to journalArticle

Gorin, Y, Leseney, AM, Ohayon, R, Dupuy, C, Pommier, J, Virion, A & Deme, D 1997, 'Regulation of the thyroid NADPH-dependent H2O2 generator by Ca2+: Studies with phenylarsine oxide in thyroid plasma membrane', Biochemical Journal, vol. 321, no. 2, pp. 383-388.
Gorin, Y. ; Leseney, A. M. ; Ohayon, R. ; Dupuy, C. ; Pommier, J. ; Virion, A. ; Deme, D. / Regulation of the thyroid NADPH-dependent H2O2 generator by Ca2+ : Studies with phenylarsine oxide in thyroid plasma membrane. In: Biochemical Journal. 1997 ; Vol. 321, No. 2. pp. 383-388.
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