Characterization of non-covalent oligomers of proteins treated with hypochlorous acid

被引:77
作者
Chapman, ALP
Winterbourn, CC
Brennan, SO
Jordan, TW
Kettle, AJ
机构
[1] Christchurch Sch Med & Hlth Sci, Dept Pathol, Free Rad Res Grp, Christchurch, New Zealand
[2] Christchurch Hosp, Mol Pathol Lab, Christchurch, New Zealand
[3] Univ Victoria, Sch Biol Sci, Wellington, New Zealand
关键词
haemoglobin; myeloperoxidase; myoglobin; neutrophil; protein aggregation; protein oxidation;
D O I
10.1042/BJ20030685
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Hypochlorous acid (HOCl) is a potent oxidant produced by myeloperoxidase that causes aggregation of many proteins. Treatment of apohaemoglobin and apomyoglobin with HOC] produced a regular series of oligomer bands when the proteins were separated by SDS/PAGE under reducing conditions. Aggregation was detectable at a HOCl/protein molar ratio of 0.5:1 and was maximal at ratios of 10: 1-20: 1. Dimers formed within 1 min of adding HOCI, and further aggregation occurred over the next 30 min. No convincing evidence for covalent cross-linking was obtained by amino acid analysis, peptide analysis or electrospray ionization-MS of HOCI-modified apomyoglobin. The latter showed an increase in mass consistent with conversion of the two methionine residues into sulphoxides. A 5-fold excess of HOCI generated approximately three chloramines on the apomyoglobin. These underwent slow decay. Protein carboryls were formed and were almost entirely located only on the polymer bands. Conversion of positively into negatively charged groups on the protein by succinylation caused preformed aggregates to dissociate. Treatment of apomyoglobin with taurine chloramine generated methionine sulphoxides but few protein carbonyls, and did not result in aggregation. We conclude that aggregation was due to strong, non-covalent interactions between protein chains. We propose that formation of protein carboryls and possibly chloramines, along with methionine oxidation, alters protein folding to expose hydrophobic areas on neighbouring molecules that associate to form dimers and higher-molecular-mass aggregates. This process could lead to the formation of aggregated proteins at sites of myeloperoxidase activity and contribute to inflammatory tissue injury.
引用
收藏
页码:33 / 40
页数:8
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