Revisit the E2 Domain of Amyloid Precursor Protein: Ferroxidase, Superoxide and Peroxynitrite Scavenging Activities
Revisit the E2 Domain of Amyloid Precursor Protein: Ferroxidase, Superoxide and Peroxynitrite Scavenging Activities
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DOI：
10.1021/acs.inorgchem.3c01336
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发表时间：
2023-06-27
影响因子：
4.6
通讯作者：
Tian，Shiliang
中科院分区：
文献类型：
--
作者：
Poore，Andrew T. T.;Zuercher，Eli C. C.;Tian，Shiliang
淀粉样前体蛋白（APP）是β-淀粉样蛋白的生物学前体，β-淀粉样蛋白是与阿尔茨海默病（AD）相关的已知组织病理学标志。APP的功能是非常有趣的，但仍然难以捉摸。APP的胞外结构域之一，E2结构域，已被提出具有铁氧化酶活性并影响神经元铁稳态。然而，报告的证据相互矛盾，其确切作用仍无定论。在这里，我们研究了铜结合位点的E2域使用扩展的X射线吸收精细结构（EXAFS），紫外-可见光，和电子顺磁共振（EPR），并发现一个新的不稳定的水配体坐标的Cu（II）的辅因子，除了四个已知的组氨酸。我们通过与亚铁的反应探索了Cu（II）-E2结构域的铁氧化酶活性，并观察到单次翻转亚铁氧化活性，速率高达1.0 × 102 M-1 s-1。Cu（I）-E2与分子氧的反应速率仅为5.3 M-1 s-1，这将限制任何潜在的多周转铁氧化酶活性到这种缓慢的速率，并阻止在多周转条件下观察活性。蛋白质的正静电势表面表明可能与带负电荷的小底物反应，如超氧自由基（O2·-）和过氧亚硝酸根（ONOO-），它们是细胞外环境中普遍存在的氧化应激的主要贡献者。结果表明，Cu（I）-E2对O2·-的去除速率为1.6 × 105 M ~（-1）s ~（-1），低于天然SOD的去除速率。然而，Cu（I）-E2与ONOO-之间的反应速率达到1.1 × 105 M-1 s-1，与天然ONOO-清除剂peroxiredoxins（105- 107 M-1 s-1）相当。因此，APP的E2结构域可以作为酶促位点，在底物限制条件下可以作为铁氧化酶、补充性O2·-清除剂和细胞铁外排通道附近的ONOO-清除剂发挥作用，并保护神经元细胞免受活性氧（ROS）和活性氮（RNS）损伤。
Amyloid precursor protein (APP) is the biological precursor of β-amyloids, a known histopathological hallmark associated with Alzheimer’s disease (AD). The function of APP is of great interest yet remains elusive. One of the extracellular domains of APP, the E2 domain, has been proposed to possess ferroxidase activity and affect neuronal iron homeostasis. However, contradicting evidence has been reported, and its precise role remains inconclusive. Here, we studied the Cu-binding site of the E2 domain using extended X-ray absorption fine structure (EXAFS), UV–vis, and electron paramagnetic resonance (EPR) and discovered that a new labile water ligand coordinates to the Cu(II) cofactor in addition to the four known histidines. We explored the proposed ferroxidase activity of the Cu(II)-E2 domain through reactions with ferrous iron and observed single-turnover ferrous oxidation activity with a rate up to 1.0 × 102M–1s–1. Cu(I)-E2 reacted with molecular oxygen at a rate of only 5.3 M–1s–1, which would restrict any potential multiturnover ferroxidase activity to this slow rate and prevents observation of activity under multiturnover conditions. The positive electrostatic potential surface of the protein indicates possible reactivity with negatively charged small substrates such as superoxide radicals (O2•–) and peroxynitrite (ONOO–) that are major contributors to the oxidative stress prevalent in the extracellular environment. Our assays showed that Cu(I)-E2 can remove O2•–at a rate of 1.6 × 105M–1s–1, which is slower than the rates of native SODs. However, the reaction between Cu(I)-E2 and ONOO–achieved a rate of 1.1 × 105M–1s–1, comparable to native ONOO–scavenger peroxiredoxins (105–107M–1s–1). Therefore, the E2 domain of APP can serve as an enzymatic site that may function as a ferroxidase under substrate-limiting conditions, a supplemental O2•–scavenger, and an ONOO–remover in the vicinity of the cellular iron efflux channel and protect neuron cells from reactive oxygen species (ROS) and reactive nitrogen species (RNS) damage.