Dithionite Reduction Kinetics of the Dissimilatory Copper-containing Nitrite Reductase of Alcalegenes xylosoxidans
Dithionite Reduction Kinetics of the Dissimilatory Copper-containing Nitrite Reductase of Alcalegenes xylosoxidans
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木糖氧化产碱菌异化含铜亚硝酸盐还原酶的连二亚硫酸盐还原动力学
DOI：
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发表时间：
2002
影响因子：
4.8
通讯作者：
R. Eady
中科院分区：
文献类型：
--
作者：
F. Yousafzai;R. Eady
我们在这里报告的第一个详细的研究连二亚硫酸盐还原动力学的含铜异化亚硝酸盐还原酶（NiR）。蓝色1型铜（T1 Cu）中心的NiR制剂，包含1型和2型铜原子的还原，遵循双相动力学。相比之下，缺乏2型铜的NiR（T2 DNiR）遵循双相动力学，二级速率常数T2 D k = 3.06 × 106 m − 1 s −1。在所有情况下，有效的还原剂都是SO 2-自由基，而不是S2 O 4 2-。所观察到的动力学是兼容的反应机制，其中T1铜的满载蛋白质直接减少连二亚硫酸盐和间接的2型铜（T2铜）网站通过分子内电子转移。T2 Cu的还原动力学与SO 2首先结合到T2 Cu中心，然后转移电子（112 s−1）将其还原一致。由于SO 2是NiR底物NO 2 −的同系物，因此它不太可能结合到催化T2 Cu位点。讨论了连二亚硫酸钠作为还原剂对酶催化活性的影响。半还原态T1 Cu（I）T2 Cu（II）的还原遵循二级动力学（k 2 = 3.33 × 107 m −1 s − 1）或一级动力学（52.6 s−1 <T1 red k 1 < 112 s−1）。T1 Cu（II）T2 Cu（II）-SO2-和T1 Cu（I）T2 Cu（II）-SO2-加合物的形成常数表明，T1 Cu的氧化还原状态影响SO2-在催化T2 Cu中心的结合.动力学分析需要开发一种数学协议，该协议可以应用于具有两个相互通信的站点的系统，但其中只有一个可以被监测。这一新的协议，首次报道，是普遍的应用。
We report here the first detailed study of the dithionite reduction kinetics of a copper-containing dissimilatory nitrite reductase (NiR). The reduction of the blue type 1 copper (T1Cu) center of NiR preparations that contained both type 1 and type 2 copper atoms, followed biphasic kinetics. In contrast, NiR that was deficient in type 2 copper (T2DNiR), followed monophasic kinetics with a second-order rate constant T2D k = 3.06 × 106 m −1s−1. In all cases the SO 2 ⨪ radical rather thanS2O 4 2 − was the effective reductant. The observed kinetics were compatible with a reaction mechanism in which the T1Cu of the fully loaded protein is reduced both directly by dithionite and indirectly by the type 2 Cu (T2Cu) site via intramolecular electron transfer. Reduction kinetics of the T2Cu were consistent with SO 2 ⨪ binding first to the T2Cu center and then transferring electrons (112 s−1) to reduce it. As SO 2 ⨪ is a homologue of NO 2 − , the NiR substrate, it is not unlikely that it binds to the catalytic T2Cu site. Effects on the catalytic activity of the enzyme using dithionite as a reducing agent are discussed. Reduction of the semireduced T1Cu(I)T2Cu(II) state followed either second-order kinetics withk 2 = 3.33 × 107 m −1 s −1or first-order kinetics with 52.6 s−1 <T1red k 1 < 112 s−1. Values of formation constants of the T1Cu(II)T2Cu(II)-SO 2 ⨪ and T1Cu(I)T2Cu(II)-SO 2 ⨪ adducts showed that the redox state of T1Cu affected binding of SO 2 ⨪ at the catalytic T2Cu center. Analysis of the kinetics required the development of a mathematical protocol that could be applied to a system with two intercommunicating sites but only one of which can be monitored. This novel protocol, reported for the first time, is of general application.
来自 Q 带 ENDOR 的关于野生型和突变型含铜亚硝酸还原酶的 1 型和 2 型铜配体环境的电子结构信息。
DOI：
10.1021/bi971604r
发表时间：
1998
期刊：
Biochemistry.
影响因子：
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作者：
Veselov,A;Olesen,K;Sienkiewicz,A;Shapleigh,JP;Scholes,CP
通讯作者：
Scholes,CP
粪产碱菌亚硝酸还原酶的结构和含锌的铜位点突变体 M150E。
影响因子：
2.9
作者：
Murphy,ME;Turley,S;Kukimoto,M;Nishiyama,M;Horinouchi,S;Sasaki,H;Tanokura,M;Adman,ET
通讯作者：
Adman,ET
影响因子：
56.9
作者：
GODDEN, JW;TURLEY, S;LEGALL, J
通讯作者：
LEGALL, J