Peroxidation of linoleate at physiological pH: Hemichrome formation by substrate binding protects against metmyoglobin activation by hydrogen peroxide

Caroline P. Baron, Leif H. Skibsted, Henrik J. Andersen

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

Peroxidation by metmyoglobin, MbFe(III), by metmyoglobin/hydrogen peroxide, MbFe(III)/H2O2, to yield the myoglobin ferryl radical ((.)MbFe(IV)=O), or by ferrylmyoglobin, MbFe(IV)=O, was investigated at physiological pH (7.4) in oil-in-water linoleate emulsions. Linoleate peroxidation was followed using second derivative ultraviolet (UV)-spectroscopy for monitoring formation of conjugated dienes and quantitative determination of specific Linoleate hydroperoxides by liquid chromatography with photodiode absorption detection. Modifications of myoglobins during lipid peroxidation were followed simultaneously by changes in the Sorer absorption band (410 or 424 nm), and in the visible absorption region (from 450 to 700 nm), combined with electron spin resonance (ESR spectroscopy for direct detection of changes in the spin state of the iron center. In contrast to MbFe(IV)=O, MbFe(III) and MbFe(III)/H2O2 were not able to initiate linoleate peroxidation in oil-in-water emulsions, and MbFe(III) was converted, by binding of linoleate (but not methyl linoleate), to a low-spin hemichrome derivate, HMbFe(III), with the distal histidine reversibly bound to the iron center. HMbFe(III) is ineffective in initiating lipid peroxidation and cannot be activated to (.)MbFe(IV)= O or MbFe(IV)=O by addition of moderate amounts of H2O2. Addition of MbFe(III) to linoleate emulsions containing H2O2 results in the competitive formation of (.)MbFe(IV)=O and HMbFe(III) in favor of HMbFe(III), and little linoleate peroxidation is detected, demonstrating the inherent protection, at physiologic pH, against peroxidation by reversible binding of the substrate to the potential myoglobin catalyst. (C) 2000 Elsevier Science Inc.
Original languageEnglish
JournalFree Radical Biology & Medicine
Volume28
Issue number4
Pages (from-to)549-558
Number of pages10
ISSN0891-5849
DOIs
Publication statusPublished - 2000
Externally publishedYes

Keywords

  • Animals
  • Emulsions
  • Horses
  • Hydrogen Peroxide
  • Hydrogen-Ion Concentration
  • Linoleic Acid
  • Lipid Peroxidation
  • Lipid Peroxides
  • Metmyoglobin
  • Spectrophotometry
  • ferrylmyoglobin
  • 12772-23-5 Metmyoglobin
  • 9KJL21T0QJ Linoleic Acid
  • BBX060AN9V Hydrogen Peroxide
  • free radical
  • heme
  • hydrogen peroxide
  • linoleic acid
  • lipid hydroperoxide
  • metmyoglobin
  • myoglobin
  • article
  • electron spin resonance
  • emulsion
  • high performance liquid chromatography
  • lipid peroxidation
  • oxidation
  • priority journal
  • protein degradation
  • protein modification
  • Ferrylmyoglobin
  • Free radicals
  • Heme-protein degradation
  • Hemichrome
  • Lipid peroxidation
  • Myoglobin Ferryl radical
  • BIOCHEMISTRY
  • ENDOCRINOLOGY
  • FATTY-ACIDS
  • LIPID-PEROXIDATION
  • FERRYL-MYOGLOBIN
  • AQUEOUS-SOLUTION
  • OXIDATION
  • REDUCTION
  • FERRYLMYOGLOBIN
  • HEMOGLOBIN
  • KINETICS
  • RADICALS
  • myoglobin ferryl radical
  • heme-protein degradation
  • hemichrome
  • free radicals
  • physiological pH
  • substrate binding
  • hemichrome formation
  • hydrogen peroxide 7722-84-1
  • linoleate 1509-85-9 peroxidation
  • metmyoglobin activation protection
  • potential myoglobin catalyst reversible substrate binding
  • 10060, Biochemistry studies - General
  • 10802, Enzymes - General and comparative studies: coenzymes
  • Biochemistry and Molecular Biophysics

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