Oxidative fragmentation of collagen and prolyl peptide by Cu(II)/H2O2. Conversion of proline residue to 2-pyrrolidone

J Biol Chem. 1992 Nov 25;267(33):23646-51.

Abstract

Oxidative degradation of collagen and the model peptides by Cu(II)/H2O2 has been studied. The depolymerization of collagen was predominantly observed by use of gel filtration chromatography. Polyproline was used as a model for collagen, and the oxidative modification was examined by amino acid analysis. Glutamic acid and gamma-aminobutyric acid were identified in the hydrolysates of oxidized polyproline. The formation of glutamic acid was reduced by treatment with NaBH4. The model peptide, (Pro-Pro-Gly)10, was also degraded by Cu(II)/H2O2, and a new N-terminal glycine was generated in proportion to the reaction time. Hydroxyl radical scavengers show only partial inhibition of the degradation of (Pro-Pro-Gly)10. In order to estimate the fragmentation mechanism, we used N-tert-butoxycarbonyl (Boc)-L-prolylglycine as a model for collagen and (Pro-Pro-Gly)10. The degradation products were isolated and characterized. Then N-tert-Boc-2-pyrrolidone, which provides gamma-aminobutyric acid by acid hydrolysis, was identified. The formation of a 2-pyrrolidone compound from oxidized Boc-L-prolylglycine is direct evidence for the scission of the peptide bond. The time-dependent formation of N-tert-Boc-2-pyrrolidone and liberation of glycine from N-tert-Boc-L-prolylglycine exposed to Cu(II)/H2O2 was observed. These results suggest that the cleavage of the peptide bond (Pro-Gly) was caused by oxidation of the proline residue, which led to the formation of the 2-pyrrolidone compound. We confirmed that proline oxidation leads to the fragmentation of proteins, accompanied by the formation of a 2-pyrrolidone structure.

MeSH terms

  • Amino Acid Sequence
  • Borohydrides
  • Chromatography, High Pressure Liquid
  • Collagen / chemistry*
  • Copper
  • Female
  • Humans
  • Hydrogen Peroxide
  • Indicators and Reagents
  • Kinetics
  • Mannitol
  • Molecular Sequence Data
  • Peptides / chemistry*
  • Peptides / isolation & purification
  • Placenta
  • Pregnancy
  • Proline*
  • Pyrrolidinones
  • Time Factors

Substances

  • Borohydrides
  • Indicators and Reagents
  • Peptides
  • Pyrrolidinones
  • polyproline
  • Mannitol
  • Copper
  • Collagen
  • Proline
  • Hydrogen Peroxide
  • 2-pyrrolidone