EPR Detection of Glutathiyl and Hemoglobin-cysteinyl Radicals during the Interaction of Peroxynitrite with Human Erythrocytes
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- 作者：Ohara Augusto ; Silvia Lopes de Menezes ; Edlaine Linares ; Natá ; lia Romero ; Rafael Radi ; and Ana Denicola
- 刊名：Biochemistry
- 出版年：2002
- 出版时间：December 3, 2002
- 年：2002
- 卷：41
- 期：48
- 页码：14323 - 14328
- 全文大小：77K
- 年卷期：v.41,no.48(December 3, 2002)
- ISSN：1520-4995
文摘
Peroxynitrite, which is formed by the fast reaction between nitric oxide and superoxide anion,has been receiving increasing attention as a mediator of human diseases. An initial controversy about thepossibility of free radical production from peroxynitrite in test tubes has been resolved, and presently itis important to establish whether peroxynitrite produces radicals in cells. Here we employed the EPRspin trapping methodology with 5,5-dimethylpyrroline N-oxide (DMPO) to study the interaction ofperoxynitrite with human erythrocytes. The results confirmed previous findings in demonstrating thatoxyhemoglobin is the main target of peroxynitrite in erythrocytes. As we first show here, the producedferryl-hemoglobin oxidizes its own amino acids and, most probably, amino acids from other hemoglobinmonomers to produce hemoglobin-tyrosyl and hemoglobin-cysteinyl radicals. In parallel, ferryl-hemoglobinalso oxidizes intracellular glutathione to produce the glutathiyl radical. The EPR spectrum of both DMPO/
cysteinyl-hemoglobin (a
H = 15.4 G) and DMPO/tyrosyl-hemoglobin (aH = 8.8 G) radical adductswas characterized. It is proposed that erythrocytes can be efficient peroxynitrite scavengers in vivo throughthe coupled action of oxyhemoglobin and glutathione. Overall, the results indicate that, through theintermediacy of carbon dioxide and/or hemoproteins, oxidation of glutathione to the glutathiyl radical islikely to be an important consequence of peroxynitrite production in vivo.
cysteinyl-hemoglobin (aH = 15.4 G) and DMPO/tyrosyl-hemoglobin (aH = 8.8 G) radical adductswas characterized. It is proposed that erythrocytes can be efficient peroxynitrite scavengers in vivo throughthe coupled action of oxyhemoglobin and glutathione. Overall, the results indicate that, through theintermediacy of carbon dioxide and/or hemoproteins, oxidation of glutathione to the glutathiyl radical islikely to be an important consequence of peroxynitrite production in vivo.