Tyrosine modifications and inactivation of active site manganese superoxide dismutase mutant (Y34F) by peroxynitrite
Tyrosine modifications and inactivation of active site manganese superoxide dismutase mutant (Y34F) by peroxynitrite
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DOI：
10.1006/abbi.1999.1202
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发表时间：
1999-06-01
影响因子：
3.9
通讯作者：
Thompson, JA
中科院分区：
文献类型：
--
作者：
MacMillan-Crow, LA;Thompson, JA
该实验室最近的研究表明，人锰超氧化物歧化酶(MnSOD)是几种慢性炎症性疾病的酪氨酸硝化靶点，包括慢性器官排斥反应、关节炎和肿瘤发生。此外，我们还证明了过氧亚硝酸盐(ONOO-)是已知的唯一一种生物氧化剂，能够灭活MnSOD中的酶活性、硝酸盐关键酪氨酸残基和诱导二叔丁氨酸的形成。为了阐明酪氨酸硝化和氧化在酶失活过程中的不同作用，我们现在比较了天然重组人MnSOD的ONOO处理(WT-MnSOD)和突变的Y34F-MnSOD，在Y34F-MnSOD中，酪氨酸34(最容易被ONOO介导的硝化作用的残基)突变为苯丙氨酸。WT-MnSOD(IC50=65 mU M，15 mU M MnSOD)和Y34F-MnSOD(IC50=55 mU M，15 mU M Y34F)对ONOO介导的灭活具有相似的剂量依赖敏感性。与WT-MnSOD相比，经ONOO-处理后，Y34F-MnSOD突变体的酪氨酸硝化效率显著降低，而二叔丁氨酸的形成显著增加。总而言之，这些结果表明，ONOO-对MnSOD的完全失活与活性部位酪氨酸残基无关，不仅包括关键酪氨酸残基的硝化，还包括酪氨酸氧化和随后形成的二酪氨酸。(C)1999年学术出版社。
Recent studies from this laboratory have demonstrated that human manganese superoxide dismutase (MnSOD) is a target for tyrosine nitration in several chronic inflammatory diseases including chronic organ rejection, arthritis, and tumorigenesis. Furthermore, we demonstrated that peroxynitrite (ONOO-) is the only known biological oxidant competent to inactivate enzymatic activity, nitrate critical tyrosine residues, and induce dityrosine formation in MnSOD. To elucidate the differential contributions of tyrosine nitration and oxidation during enzymatic inactivation, we now compare ONOO- treatment of native recombinant human MnSOD (WT-MnSOD) and a mutant, Y34F-MnSOD, in which tyrosine 34 (the residue most susceptible to ONOO--mediated nitration) was mutated to phenylalanine. Both WT-MnSOD (IC50 = 65 mu M, 15 mu M MnSOD) and Y34F-MnSOD (IC50 = 55 mu M, 15 mu M Y34F) displayed similar dose-dependent sensitivity to ONOO--mediated inactivation. Compared to WT-MnSOD, the Y34F-MnSOD mutant demonstrated significantly less efficient tyrosine nitration and enhanced formation of dityrosine following treatment with ONOO-. Collectively, these results suggest that complete inactivation of MnSOD by ONOO- can occur independent of the active site tyrosine residue and includes not only nitration of critical tyrosine residues but also tyrosine oxidation and subsequent formation Of dityrosine. (C) 1999 Academic Press.