Peroxynitrite-mediated oxidation of plasma fibronectin
Peroxynitrite-mediated oxidation of plasma fibronectin
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DOI：
10.1016/j.freeradbiomed.2016.06.013
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发表时间：
2016-08-01
影响因子：
7.4
通讯作者：
Davies, Michael J.
中科院分区：
文献类型：
--
作者：
Degendorfer, Georg;Chuang, Christine Y.;Davies, Michael J.
纤连蛋白是存在于人血浆和基底膜中的大的二聚体糖蛋白。后者是动脉壁中内皮细胞下面的特化细胞外基质。过氧亚硝酸（ONOOH）是一种有效的氧化剂和硝化剂，在体内由超氧化物和一氧化氮自由基通过刺激巨噬细胞和其他细胞形成。相当多的证据支持ONOOH参与人类动脉粥样硬化病变的发展和破裂，可能通过细胞外基质损伤。在这里，我们证明了人血浆纤连蛋白上的Tyr和Trp残基对ONOOH高度敏感，这导致形成3-硝基酪氨酸、6-硝基色氨酸和二酪氨酸以及蛋白质聚集和片段化。这发生在等摩尔或更高水平的氧化剂，并以剂量依赖性的方式。在占母体氨基酸丰度后，Tyr的修饰在数量上比Trp更显著（9.1%对1.5%转化率，500 μ M ONOOH），但仅占添加的总氧化剂的一小部分。LC-MS研究鉴定了28个硝化位点（24个Tyr，4个Trp），其中许多存在于对蛋白质功能至关重要的结构域中，包括细胞结合和anastellin结构域。与对照组相比，人冠状动脉内皮细胞在ONOOH修饰的纤连蛋白上的粘附性和细胞铺展性降低，这与修饰基质诱导的细胞功能障碍一致。对人类动脉粥样硬化病变的研究为3-硝基酪氨酸和纤连蛋白的共定位提供了证据。ONOOH介导的纤连蛋白修饰和受损的细胞-基质相互作用可能导致内皮细胞功能障碍，动脉粥样硬化病变的纤维帽减弱，以及破裂倾向增加。(C)2016爱思唯尔B. V.保留所有权利。
Fibronectin is a large dimeric glycoprotein present in both human plasma and in basement membranes. The latter are specialized extracellular matrices underlying endothelial cells in the artery wall. Peroxynitrous acid (ONOOH) a potent oxidizing and nitrating agent, is formed in vivo from superoxide and nitric oxide radicals by stimulated macrophages and other cells. Considerable evidence supports ONOOH involvement in human atherosclerotic lesion development and rupture, possibly via extracellular matrix damage. Here we demonstrate that Tyr and Trp residues on human plasma fibronectin are highly sensitive to ONOOH with this resulting in the formation of 3-nitrotyrosine, 6-nitrotryptophan and dityrosine as well as protein aggregation and fragmentation. This occurs with equimolar or greater levels of oxidant, and in a dose-dependent manner. Modification of Tyr was quantitatively more significant than Trp (9.1% versus 1.5% conversion with 500 mu M ONOOH) after accounting for parent amino acid abundance, but only accounts for a small percentage of the total oxidant added. LC-MS studies identified 28 nitration sites (24 Tyr, 4 Trp) with many of these present within domains critical to protein function, including the cell-binding and anastellin domains. Human coronary artery endothelial cells showed decreased adherence and cell-spreading on ONOOH-modified fibronectin compared to control, consistent with cellular dysfunction induced by the modified matrix. Studies on human atherosclerotic lesions have provided evidence for co-localization of 3-nitrotyrosine and fibronectin. ONOOH-mediated fibronectin modification and compromised cell-matrix interactions, may contribute to endothelial cell dysfunction, a weakening of the fibrous cap of atherosclerotic lesions, and an increased propensity to rupture. (C) 2016 Elsevier B.V. All rights reserved.