Intraprotein electron transfer between the FMN and heme domains in endothelial nitric oxide synthase holoenzyme
Intraprotein electron transfer between the FMN and heme domains in endothelial nitric oxide synthase holoenzyme
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DOI：
10.1016/j.bbapap.2011.08.004
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发表时间：
2011-12-01
影响因子：
3.2
通讯作者：
Tollin, Gordon
中科院分区：
文献类型：
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作者：
Feng, Changjian;Taiakina, Valentina;Tollin, Gordon
从黄素单核苷酸 (FMN) 到血红素的蛋白内电子转移 (IET) 是一氧化氮合酶 (NOS) 合成一氧化氮 (NO) 的重要步骤。神经元和诱导型 NOS（nNOS 和 iNOS）全酶的 IET 动力学先前已在我们的实验室中通过激光闪光光解作用确定[综述于：C.J. Feng, G. Tollin, Dalton Trans., (2009) 6692-6700]。在此，我们报告了在添加钙调蛋白 (CaM) 的情况下，牛内皮 NOS (eNOS) 全酶中 IET 的动力学。 CaM 存在下的 IET 速率常数估计类似于 4.3 s(-1)。在没有 CaM 的情况下没有观察到 IET，这表明 CaM 是控制 eNOS 酶中 FMN - 血红素 IET 的主要因素。 eNOS 全酶的 IET 速率常数值比 iNOS 和 CaM 结合的 nNOS 全酶获得的 IET 速率常数值大约小 10 倍。讨论了 NOS 异构体之间 JET 动力学差异的可能机制。由于这些酶的 IET 过程中的限速步骤是从输入状态到输出状态的构象变化，因此较慢的构象变化（比其他亚型）最有可能导致 eNOS 中的 IET 较慢。 (C) 2011 Elsevier B.V. 保留所有权利。
Intraprotein electron transfer (IET) from Flavin mononucleotide (FMN) to heme is an essential step in nitric oxide (NO) synthesis by NO synthase (NOS). The IET kinetics in neuronal and inducible NOS (nNOS and iNOS) holoenzymes have been previously determined in our laboratories by laser flash photolysis [reviewed in: C.J. Feng, G. Tollin, Dalton Trans., (2009) 6692-6700]. Here we report the kinetics of the IET in a bovine endothelial NOS (eNOS) holoenzyme in the presence and absence of added calmodulin (CaM). The IET rate constant in the presence of CaM is estimated to be similar to 4.3 s(-1). No IET was observed in the absence of CaM, indicating that CaM is the primary factor in controlling the FMN - heme IET in the eNOS enzyme. The IET rate constant value for the eNOS holoenzyme is approximately 10 times smaller than those obtained for the iNOS and CaM-bound nNOS holoenzymes. Possible mechanisms underlying the difference in JET kinetics among the NOS isoforms are discussed. Because the rate-limiting step in the IET process in these enzymes is the conformational change from input state to output state, a slower conformational change (than in the other isoforms) is most likely to cause the slower IET in eNOS. (C) 2011 Elsevier B.V. All rights reserved.