Radical-induced chain oxidation of proteins and its inhibition by chain-breaking antioxidants.

Exposure of proteins to oxygen-centred radicals results in dramatic changes in their structure, stability and function; properties that have been studied in many laboratories from a qualitative viewpoint. To allow a quantitative evaluation, we subjected aerated solutions of BSA to hydroxyl and superoxide anion radicals generated radiolytically under conditions where all radicals formed reacted with the protein (as judged by maximum damage to BSA). We observed that for each radical generated approx. 15 amino acids were consumed initially. Similar results were found with lysozyme and melittin. Such a massive consumption of BSA's amino acids was not observed when irradiations were carried out under anaerobic conditions. When bilirubin or Trolox (a water-soluble analogue of vitamin E) was added at a 2-fold molar excess over BSA, initial consumption of all measured amino acids, except tryptophan, decreased 4-fold. The total mass of amino acids initially protected (from consumption) exceeded the mass of antioxidants consumed by more than a factor of 20. Such protection of amino acids was not observed when the antioxidant-inactive acetyl Trolox was used. These results suggest that radical-mediated oxidation of proteins can proceed via a previously unrecognized chain reaction that may be inhibited by chain-breaking antioxidants.