Characterization of the adduct formed from the reaction between homocysteine thiolactone and low-density lipoprotein: Antioxidant implications

Eric Ferguson, Neil Hogg, William E. Antholine, Joy Joseph, Ravinder Jit Singh, Sampath Parthasarathy, B. Kalyanaraman

Research output: Contribution to journalArticle

32 Scopus citations

Abstract

Homocysteine thiolactone is a cyclic thioester that is implicated in the development of atherosclerosis. This molecule will readily acylate primary amines, forming a homocystamide adduct, which contains a primary amine and a thiol. Here, we have characterized and evaluated the antioxidant potential of the homocystamide-low-density lipoprotein (LDL) adduct, a product of the reaction between homocysteine thiolactone and LDL. Treatment of LDL with homocysteine thiolactone resulted in a time-dependent increase in LDL-bound thiols that reached approximately 250 nmol thiol/mg LDL protein. The thiol groups of the homocystamide-LDL adduct were labeled with the thiol-reactive nitroxide, methanethiosulfonate spin label. Using paramagnetic relaxing agents and the electron spin resonance spin labeling technique, we determined that the homocystamide adducts were predominately exposed to the aqueous phase. The homocystamide-LDL adduct was resistant to myoglobin- and Cu2+- mediated oxidation (with respect to native LDL), as measured by the formation of conjugated dienes and thiobarbituric acid reactive substances, and the depletion of vitamin E. This antioxidant effect was due to increased thiol content, as the effect was abolished with N-ethylmaleamide pre-treatment. We conclude that the reaction between homocysteine thiolactone and LDL generates an LDL molecule that is more resistant to oxidative modification than native LDL. The potential relationship between the homocystamide-LDL adduct and the development of atherosclerosis is discussed.

Original languageEnglish (US)
Pages (from-to)968-977
Number of pages10
JournalFree Radical Biology and Medicine
Volume26
Issue number7-8
DOIs
StatePublished - Apr 1 1999

Keywords

  • Antioxidants
  • Electron spin resonance
  • Homocystamide adduct
  • Homocysteine
  • Homocysteine thiolactone
  • Lipid peroxidation
  • Low-density lipoprotein
  • Protein thiols

ASJC Scopus subject areas

  • Biochemistry
  • Physiology (medical)

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