Effects of polyaminocarboxylate metal chelators on iron-thiolate induced oxidation of methionine- and histidine-containing peptides.
OBJECTIVE
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 (EDDA), and ethylenediaminetetraacetate (EDTA), but they were slowed down by ethyl-enebis(oxyethylenenitrilo)tetraacetate (EGTA) and diethylenetri-aminepentaacetate (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.
Zhao
F
F
Department of Pharmaceutical Chemistry, University of Kansas, Lawrence 66047, USA. off
Yang
J
J
Schöneich
C
C
D000641:Ammonia / Q000737:chemistry; D002264:Carboxylic Acids / Q000737:chemistry*; D002374:Catalase / Q000737:chemistry; D002614:Chelating Agents / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D005290:Ferric Compounds / Q000737:chemistry*; D016166:Free Radical Scavengers / Q000737:chemistry; D006639:Histidine / Q000737:chemistry*; D007502:Iron Chelating Agents / Q000737:chemistry*; D008715:Methionine / Q000737:chemistry*; D009842:Oligopeptides / Q000737:chemistry*; D010084:Oxidation-Reduction; D017382:Reactive Oxygen Species; D013454:Sulfoxides / Q000737:chemistry
D002264:Carboxylic Acids; D002614:Chelating Agents; D005290:Ferric Compounds; D016166:Free Radical Scavengers; D007502:Iron Chelating Agents; D009842:Oligopeptides; D017382:Reactive Oxygen Species; D013454:Sulfoxides; D006639:Histidine; D000641:Ammonia; D008715:Methionine; D002374:Catalase