Nitric Oxide Modulates Endonuclease III Redox Activity by a 800 mV Negative Shift upon [Fe(4)S(4)] Cluster Nitrosylation.
Nitric Oxide Modulates Endonuclease III Redox Activity by a 800 mV Negative Shift upon [Fe(4)S(4)] Cluster Nitrosylation.
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DOI：
10.1021/jacs.8b07362
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发表时间：
2018-09-19
影响因子：
15
通讯作者：
Barton JK
中科院分区：
文献类型：
--
作者：
Ekanger LA;Oyala PH;Moradian A;Sweredoski MJ;Barton JK
在这里，我们的特点[Fe 4S 4]簇亚硝基化的DNA修复酶，核酸内切酶III（EndoIII），使用DNA修饰的金电化学和蛋白质膜伏安法，电泳迁移率变动分析，质谱的整个和胰蛋白酶消化的蛋白质，和各种光谱。在厌氧条件下，EndoIII暴露于一氧化氮时，[Fe 4S 4]簇以1：1的比例转化为二亚硝酰铁络合物[（Cys）2 Fe（NO）2]-和红曲霉素红酯[（μ-Cys）2Fe 2（NO）4]，每个亚硝酰化簇平均保留3.05 ± 0.01 Fe。二亚硝酰铁络合物的形成与以前的报道一致，但黄曲霉素红酯是EndoIII亚硝酰化的未报道产物。超精细次能级相关（HYSCORE）脉冲EPR光谱检测到两个不同类别的NO与14 N超精细耦合一致的二亚硝酰铁络合物和还原的红曲霉素的红色酯。用14 NO和15 NO亚硝基化的EndoIII的全蛋白质质谱支持蛋白结合的[（μ-Cys）2Fe 2（NO）4]胡萝卜素红酯的归属。使用DNA修饰的金电化学可观察到DNA结合的EndoIII的[Fe 4S 4]2+/3+氧化还原对，但亚硝基化的EndoIII使用DNA电化学在金上不显示可观察到的氧化还原活性，尽管其具有与天然蛋白相似的DNA结合亲和力。然而，石墨上的蛋白质膜的直接电化学显示，天然和亚硝基化的EndoIII的还原电位分别为127 ± 6和−674 ± 8 mV（相对于NHE），对应于簇亚硝基化的位移约为−800 mV。总的来说，这些数据表明，DNA结合的氧化还原活性，并通过扩展DNA介导的电荷传输，调制[Fe 4S 4]簇亚硝基化。
Here we characterize the [Fe4S4] cluster nitrosylation of a DNA repair enzyme, endonuclease III (EndoIII), using DNA-modified gold electrochemistry and protein film voltammetry, electrophoretic mobility shift assays, mass spectrometry of whole and trypsin-digested protein, and a variety of spectroscopies. Exposure of EndoIII to nitric oxide under anaerobic conditions transforms the [Fe4S4] cluster into a dinitrosyl iron complex, [(Cys)2 Fe(NO)2]-, and Roussin’s red ester, [(μ-Cys)2Fe2(NO)4], in a 1:1 ratio with an average retention of 3.05 ± 0.01 Fe per nitrosylated cluster. The formation of the dinitrosyl iron complex is consistent with previous reports, but the Roussin’s red ester is an unreported product of EndoIII nitrosylation. Hyperfine sublevel correlation (HYSCORE) pulse EPR spectroscopy detects two distinct classes of NO with 14N hyperfine couplings consistent with the dinitrosyl iron complex and reduced Roussin’s red ester. Whole-protein mass spectrometry of EndoIII nitrosylated with 14NO and 15NO support the assignment of a protein-bound [(μ-Cys)2Fe2(NO)4] Roussin’s red ester. The [Fe4S4]2+/3+ redox couple of DNA-bound EndoIII is observable using DNA-modified gold electrochemistry, but nitrosylated EndoIII does not display observable redox activity using DNA electrochemistry on gold despite having a similar DNA-binding affinity as the native protein. However, direct electrochemistry of protein films on graphite reveals the reduction potential of native and nitrosylated EndoIII to be 127 ± 6 and −674 ± 8 mV vs NHE, respectively, corresponding to a shift of approximately −800 mV with cluster nitrosylation. Collectively, these data demonstrate that DNA-bound redox activity, and by extension DNA-mediated charge transport, is modulated by [Fe4S4] cluster nitrosylation.
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影响因子：
15
作者：
Grodick, Michael A.;Segal, Helen M.;Zwang, Theodore J.;Barton, Jacqueline K.
通讯作者：
Barton, Jacqueline K.
DOI：
10.1073/pnas.74.8.3203
发表时间：
1977-01-01
影响因子：
11.1
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作者：
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作者：
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