Oxidation of the zinc-thiolate complex and uncoupling of endothelial nitric oxide synthase by peroxynitrite.
Oxidation of the zinc-thiolate complex and uncoupling of endothelial nitric oxide synthase by peroxynitrite.
复制标题
DOI：
10.1172/jci14442
复制
发表时间：
2002-03
期刊：
影响因子：
--
通讯作者：
M. Zou;Chaomei Shi;R. Cohen
中科院分区：
文献类型：
--
作者：
M. Zou;Chaomei Shi;R. Cohen
一氧化氮(NO)是由许多细胞中的一氧化氮合酶(NOS)产生的，在神经、肌肉、心血管和免疫系统中发挥着重要作用。在不同的疾病条件下，所有三种类型的一氧化氮合酶(神经元型、诱导型和内皮型)都被报道通过未知的机制产生氧化剂。我们在这里提出了第一个证据，过氧亚硝酸盐(ONOO(-))从内皮一氧化氮合酶(ENOS)的锌硫酸盐簇(ENOS)中释放锌，并推测在单体之间形成二硫键。结果，在还原条件下，原本抵抗十二烷基硫酸钠的eNOS二聚体发生破坏。ENOS的催化活性对ONOO(-)非常敏感，ONOO(-)减少了NO的合成，增加了酶产生超氧阴离子(O(2)(.-))。还原辅因子四氢生物蝶呤不被氧化，也不能阻止eNOS被同样低浓度的Oono(-)氧化。此外，来自内皮细胞的eNOS暴露在高糖环境中产生更多的O(2)(.-)，并且，像从糖尿病低密度脂蛋白受体缺陷小鼠提纯的eNOS一样，含有更少的锌和更少的抗十二烷基硫酸钠二聚体。因此，内皮型一氧化氮合酶暴露于包括ONOO(-)在内的氧化剂可导致糖尿病时酶解偶联和O(2)(.-)的产生增加，从而进一步促进内皮细胞氧化应激。ONOO(-)对一氧化氮合酶锌硫酸盐中心的调节为疾病中酶功能的调节提供了新的机制。
Nitric oxide (NO) is produced by NO synthase (NOS) in many cells and plays important roles in the neuronal, muscular, cardiovascular, and immune systems. In various disease conditions, all three types of NOS (neuronal, inducible, and endothelial) are reported to generate oxidants through unknown mechanisms. We present here the first evidence that peroxynitrite (ONOO(-)) releases zinc from the zinc-thiolate cluster of endothelial NOS (eNOS) and presumably forms disulfide bonds between the monomers. As a result, disruption of the otherwise SDS-resistant eNOS dimers occurs under reducing conditions. eNOS catalytic activity is exquisitely sensitive to ONOO(-), which decreases NO synthesis and increases superoxide anion (O(2)(.-)) production by the enzyme. The reducing cofactor tetrahydrobiopterin is not oxidized, nor does it prevent oxidation of eNOS by the same low concentrations of OONO(-). Furthermore, eNOS derived from endothelial cells exposed to elevated glucose produces more O(2)(.-), and, like eNOS purified from diabetic LDL receptor-deficient mice, contains less zinc and fewer SDS-resistant dimers. Hence, eNOS exposure to oxidants including ONOO(-) causes increased enzymatic uncoupling and generation of O(2)(.-) in diabetes, contributing further to endothelial cell oxidant stress. Regulation of the zinc-thiolate center of NOS by ONOO(-) provides a novel mechanism for modulation of the enzyme function in disease.