Mechanism of lysozyme inactivation and degradation by iron

Hassan Sellak, Elisabeth Franzini, Jacques Hakim, Catherine Pasquier

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16 Scopus citations

Abstract

The site-specific lysozyme damage by iron and by iron-catalysed oxygen radicals was investigated. A solution of purified lysozyme was inactivated by Fe(II) at pH 7.4 in phosphate buffer, as tested on cleavage of Micrococcus lysodeikticus cells; this inactivation was time- and iron concentration-dependent and was associated with a loss of tryptophan fluorescence. In addition, it was reversible at pH 4, as demonstrated by lysozyme reactivation and by the intensity of the 14.4-kD-band on SDS-PAGE. Desferal (1 mm) and Detapac (1 mm) added before iron, prevented lysozyme inactivation, while catalase (100 μg/ ml), superoxide dismutase (100 μg/ml) and bovine serum albumin (100 μg/ml) gave about 30 to 40% protection by competing with lysozyme for iron binding. The denaturing effect of iron on lysozyme was studied in the presence of H2O2 (1 mm) and ascorbate (1 mm); under these conditions the enzyme underwent partly irreversible inactivation and degradation different to that produced by gamma radiolysis-generated .OH. Catalase almost fully protected lysozyme; in contrast, mannitol (10 mm), benzoate (10 mm), and formate (10 mm) provided no protection because of their inability to access the site at which damaging species are generated. In this system, radical species were formed in a site-specific manner, and they reacted essentially with lysozyme at the site of their formation, causing inactivation and degradation differently than the hydroxyl radical.

Original languageEnglish (US)
Pages (from-to)172-178
Number of pages7
JournalArchives of Biochemistry and Biophysics
Volume299
Issue number1
DOIs
StatePublished - Nov 15 1992

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology

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