Pharmaceutical Research

, Volume 13, Issue 6, pp 931–938 | Cite as

Effects of Polyaminocarboxylate Metal Chelators on Iron-thiolate Induced Oxidation of Methionine- and Histidine-Containing Peptides

  • Fang Zhao
  • Jian Yang
  • Christian Schöneich
Article

Abstract

Purpose. Site-specific protein oxidation induced by prooxidant/metal/ oxygen has been recognized as one of the major degradation pathways of protein pharmaceuticals. Polyaminocarboxylate (PAC) metal chelators are commonly employed to prevent metal-catalyzed oxidation, for they sequester metals. However, studies have indicated that iron chelates may still be catalytically active due to their specific coordination geometry. The purpose of this study was to investigate how PAC chelators affect prooxidant/metal/oxygen-catalyzed oxidation of peptides containing histidine (His) and methionine (Met).

Methods. PACs were applied to a model oxidizing system, dithiothreitol/iron/oxygen, which was shown to promote the oxidation of Met to Met sulfoxide in the two model peptides, GGGMGGG and GHGMGGG.

Results. PAC chelators did not suppress the peptide oxidation but significantly changed the product pattern. In particular, the yield of Met sulfoxide dropped significantly, while a number of other products emerged, including oxidation products from the N-terminus and His (if present). Overall, the oxidation became rather non-selective in the presence of PACs. The oxidation kinetics were significantly accelerated by nitrilotriacetate (NTA), ethylenediaminediacetate (HDDA), and ethylenediaminetetraacetate (EDTA), but they were slowed down by ethyl-enebis(oxyethylenenitrilo)tetraacetate (EGTA) and diethylenetriaminepentaacetate (DTPA). Meanwhile the PAC chelators were also observed to undergo degradation. Scavengers of hydrogen peroxide or hydroxyl radicals exerted only partial inhibition on the peptide oxidation.

Conclusions. The results of this study are rationalized by the abilities of PAC chelators (i) to extract iron from potential binding sites of the peptides to impair site-specific oxidation, and (ii) to promote the formation of ROS different from the species formed at the peptide metal-binding sites.

iron chelator oxidation methionine/histidine 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

REFERENCES

  1. 1.
    E. R. Stadtman. Oxidation of free amino acids and amino acid residues in proteins by radiolysis and by metal-catalyzed reactions. Annu. Rev. Biochem. 62:797–821 (1993).Google Scholar
  2. 2.
    E. R. Stadtman and C. N. Oliver. Metal-catalyzed oxidation of proteins. J. Biol. Chem. 266:2005–2008 (1991).Google Scholar
  3. 3.
    E. R. Stadtman. Metal-ion catalyzed oxidation of proteins: Biochemical mechanism and biological consequences. Free Rad. Biol. Med. 9:315–325 (1990).Google Scholar
  4. 4.
    M. C. Manning, K. Patel, and R. T. Borchardt. Stability of protein pharmaceuticals. Pharm. Res. 6:903–918 (1989).Google Scholar
  5. 5.
    W. Vogt. Oxidation of methionyl residues in proteins: tools, targets, and reversal. Free Rad. Biol. Med. 18:93–105 (1995).Google Scholar
  6. 6.
    S. Li, T. Nguyen, C. Schöneich, and R. T. Borchardt. Aggregation and precipitation of human relaxin induced by metal-catalyzed oxidation. Biochemistry 34:5762–5772 (1995).Google Scholar
  7. 7.
    R. Pearlman and T. Nguyen. Pharmaceutics of protein drugs. J. Pharm. Pharmacol. 44(suppl.1):178–185 (1992).Google Scholar
  8. 8.
    G. W. Becker, P. M. Tackitt, W. W. Bromer, D. S. Lefeber, and R. M. Riggin. Isolation and characterization of a sulfoxide and a desamido derivative of biosynthetic human growth hormone. Biotechnol. Appl. Biochem. 10:326–337 (1988).Google Scholar
  9. 9.
    B. C. Cunningham, M. G. Mulkerrin, J. A. Wells. Dimerization of human growth hormone by zinc. Science 253:545–548 (1991).Google Scholar
  10. 10.
    Ch. Schöneich, F. Zhao, G. S. Wilson, and R. T. Borchardt. Ironthiolate induced oxidation of methionine to methionine sulfoxide in small model peptides. Intramolecular catalysis by histidine. Biochim. Biophys. Acta 1158:307–322 (1993).Google Scholar
  11. 11.
    E. Atherton and R. C. Sheppard. Solid phase peptide synthesis. A practical approach. IRL Press at Oxford University Press. 1989.Google Scholar
  12. 12.
    G. A. Elgavish and J. Granot. Enhancement of 31P relaxation rates of orthophosphate and ATP in the presence of EDTA. Evidence for EDTA-Fe(III)-phosphate ternary complexes. J. Magn. Reson. 36:147–150 (1979).Google Scholar
  13. 13.
    M. Deacon, M. R. Smyth, and L. G. M. T. Tuinstra. Chromatographic separations of metal chelates present in commercial fertilizers. II. Development of an ion-pair chromatographic separation for the simultaneous determination of Fe(III) chelates of EDTA, DTPA, HEEDTA, EDDHA and EDDHMA, and the Cu(II), Zn(II) and Mn(II) chelates of EDTA. J. Chromatog. A 659:349–357 (1994).Google Scholar
  14. 14.
    M. S. Akhlaq and C. v. Sonntag. Free-radical-induced elimination of H2S from dithiothreitol. A chain reaction. J. Am. Chem. Soc. 108:3542–3544 (1986).Google Scholar
  15. 15.
    K. Uchida and S. Kawakishi. Selective oxidation of imidazole ring in histidine residues by the ascorbic acid-copper ion system. Biochem. Biophys. Res. Commun. 138:659–665 (1986).Google Scholar
  16. 16.
    R. L. Levine. Oxidation modification of glutamine synthase. J. Biol. Chem. 258:11823–11827 (1983).Google Scholar
  17. 17.
    R. T. Dean, S. P. Wolff, and M. A. McElligott. Histidine and proline are important sites of free radical damage to proteins. Free Rad. Res. Comms. 7:97–103 (1989).Google Scholar
  18. 18.
    W. M. Garrison. Reaction mechanisms in the radiolysis of peptides, polypeptides, and proteins. Chem. Rev. 87:381–398 (1987).Google Scholar
  19. 19.
    K. C. Francis, D. Cummins and J. Oakes. Kinetics and structural investigations of [FeIII(edta)]-[edta=ethylenediaminetetra-acetate(4-)] catalyzed decomposition of hydrogen peroxide. J. Chem. Soc. Dalton Trans. 493–501 (1985).Google Scholar
  20. 20.
    G. V. Buxton, C. L. Greenstock, W. P. Helman, and A. B. Ross. Critical review of rate constants for reactions of hydrated electrons, hydrogen atoms and hydroxyl radicals in aqueous solution. J. Phys. Chem. Ref. Data 17:513–886 (1988).Google Scholar
  21. 21.
    J. D. Rush and W. H. Koppenol. The reaction between ferrous polyaminocarboxylate complexes and hydrogen peroxide: an investigation of the reaction intermediates by stopped flow spectrophotometry. J. Inorg. Biochem. 29:199–215 (1987).Google Scholar
  22. 22.
    M. A. Miller, D. Bandyopadyay, J. M. Mauro, T. G. Traylor, and J. Kraut. Reaction of ferrous cytochrome c peroxidase with dioxygen: site-directed mutagenesis provides evidence for rapid reduction of dioxygen by intramolecular electron transfer from the compound I radical site. Biochemistry 31:1992 (1992).Google Scholar
  23. 23. (a)
    N. Zhang, H. P. Schuchmann, and C. von Sonntag. The reaction of superoxide radicalanion with dithiothreitol—a chain process. J. Phys. Chem. 95:4718–4722 (1991). (b) P. K. Sysak, C. S. Foote, and Ta-Y. Ching. Chemistry of singlet oxygen-XXV. Photooxygenation of methionine. Photochem. Photobiol. 26:19–27 (1977) (c) calculated from catalase activity defined by Sigma.Google Scholar
  24. 24.
    L. G. Sillèn. Stability constants of metal ion complexes. Chemical Society. London. 1964.Google Scholar
  25. 25.
    J. L. Hoard, M. Lind, and J. V. Silverton. The stereochemistry of the ethylenediamine-tetraacetatoaquoferrate(III) ion. J. Am. Chem. Soc. 83:2770–2771 (1961).Google Scholar
  26. 26.
    Ch. Schöneich, A. Aced, and K.-D. Asmus. Mechanism of oxidation of aliphatic thioethers to sulfoxides by hydroxyl radical. The importance of molecular oxygen. J. Am. Chem. Soc. 115:11376–11383 (1993).Google Scholar
  27. 27.
    S. Rahhal and H. W. Richter. Reduction of hydrogen peroxide by the ferrous iron chelate of diethylenetriamin-N,N,N',N″,N″-pentaacetate. J. Am. Chem. Soc. 110:3126–3133 (1988).Google Scholar
  28. 28.
    Ch. Schöneich and J. Yang. Oxidation of methionine peptides by Fenton systems: The importance of peptide sequence, neighboring groups, and EDTA. J. Chem. Soc. Perkin Trans II (1996), in press.Google Scholar
  29. 29.
    J. D. Rush and W. H. Koppenol. Reactions of FeIInta and FeII2edda with hydrogen peroxide. J. Am. Chem. Soc. 110:4957–4963 (1988).Google Scholar

Copyright information

© Plenum Publishing Corporation 1996

Authors and Affiliations

  • Fang Zhao
    • 1
  • Jian Yang
    • 1
    • 2
  • Christian Schöneich
    • 1
  1. 1.Department of Pharmaceutical ChemistryUniversity of KansasLawrence
  2. 2.Amgen Inc.Thousand Oaks

Personalised recommendations