Radical damage to proteins studied by EPR spin-trapping techniques

J. L. Clément, B. C. Gilbert, A. Rockenbauer, P. Tordo

Research output: Contribution to journalArticle

20 Citations (Scopus)

Abstract

We report the use of the novel spin trap DEPMPO in conjunction with EPR spectroscopy to provide evidence for free-radical mediated oxidation of some proteins in aqueous solution, with systems including metal ions, peroxides or both. In addition we have used a novel approach involving detailed spectral simulation (to reproduce anisotropic features of phosphorus, hydrogen and nitrogen splittings) to confirm structural assignments. Oxidation of bovine serum albumin (BSA) with either Ce4+ or IrCl62- leads to the predominant formation of spin-trapped oxygen-centred free-radicals, attributed to tyrosyl-based adducts, a reaction also observed for sulfur-free analogues (lysozyme and tyrosine-containing models). Similar spectra were obtained from reactions involving oxidation with HRP/H2O2 and methaemoglobin/H2O2 couples, through intermolecular oxidation of the proteins via oxo-iron intermediates. Oxidation of BSA with peroxynitrite leads to an anisotropic spectrum which is assigned to the trapping of a sulfur-centred radical. This assignment has been confirmed via denaturation of the spin-adduct (and simulation of the resulting less immobilized spectrum). Reaction of the hydroxyl radical with BSA and lysozyme, in Fenton-type reactions, leads to the detection of anisotropic spectra derived from the spin-trapping of carbon-centred radicals, whose assignment is discussed.

Original languageEnglish
Pages (from-to)1463-1470
Number of pages8
JournalJournal of the Chemical Society, Perkin Transactions 2
Issue number9
Publication statusPublished - 2001

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Paramagnetic resonance
Oxidation
Bovine Serum Albumin
Muramidase
Proteins
Sulfur
Free Radicals
Methemoglobin
Denaturation
Peroxynitrous Acid
Peroxides
Hydroxyl Radical
Phosphorus
Metal ions
Tyrosine
Hydrogen
Nitrogen
Carbon
Iron
Spectroscopy

ASJC Scopus subject areas

  • Chemistry(all)

Cite this

Radical damage to proteins studied by EPR spin-trapping techniques. / Clément, J. L.; Gilbert, B. C.; Rockenbauer, A.; Tordo, P.

In: Journal of the Chemical Society, Perkin Transactions 2, No. 9, 2001, p. 1463-1470.

Research output: Contribution to journalArticle

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