Yang Zhi-Yong, Badalyan Artavazd, Hoffman Brian M, Dean Dennis R, Seefeldt Lance C
Department of Chemistry and Biochemistry, Utah State University, Logan, Utah 84322, United States.
Departments of Chemistry and Molecular Biosciences, Northwestern University, Evanston, Illinois 60208, United States.
J Am Chem Soc. 2023 Mar 15;145(10):5637-5644. doi: 10.1021/jacs.2c09576. Epub 2023 Mar 1.
A central feature of the current understanding of dinitrogen (N) reduction by the enzyme nitrogenase is the proposed coupling of the hydrolysis of two ATP, forming two ADP and two Pi, to the transfer of one electron from the Fe protein component to the MoFe protein component, where substrates are reduced. A redox-active [4Fe-4S] cluster associated with the Fe protein is the agent of electron delivery, and it is well known to have a capacity to cycle between a one-electron-reduced [4Fe-4S] state and an oxidized [4Fe-4S] state. Recently, however, it has been shown that certain reducing agents can be used to further reduce the Fe protein [4Fe-4S] cluster to a super-reduced, all-ferrous [4Fe-4S] state that can be either diamagnetic ( = 0) or paramagnetic ( = 4). It has been proposed that the super-reduced state might fundamentally alter the existing model for nitrogenase energy utilization by the transfer of two electrons per Fe protein cycle linked to hydrolysis of only two ATP molecules. Here, we measure the number of ATP consumed for each electron transfer under steady-state catalysis while the Fe protein cluster is in the [4Fe-4S] state and when it is in the [4Fe-4S] state. Both oxidation states of the Fe protein are found to operate by hydrolyzing two ATP for each single-electron transfer event. Thus, regardless of its initial redox state, the Fe protein transfers only one electron at a time to the MoFe protein in a process that requires the hydrolysis of two ATP.
目前对固氮酶还原二氮（N）的理解的一个核心特征是，有人提出将两个ATP水解生成两个ADP和两个Pi，与一个电子从铁蛋白组分转移到钼铁蛋白组分相偶联，底物在钼铁蛋白组分上被还原。与铁蛋白相关的一个具有氧化还原活性的[4Fe-4S]簇是电子传递的媒介，众所周知它能够在单电子还原的[4Fe-4S]状态和氧化的[4Fe-4S]状态之间循环。然而，最近有研究表明，某些还原剂可用于将铁蛋白的[4Fe-4S]簇进一步还原为超还原的全亚铁[4Fe-4S]状态，该状态可能是抗磁性的（ = 0）或顺磁性的（ = 4）。有人提出，超还原状态可能会从根本上改变现有的固氮酶能量利用模型，即每个铁蛋白循环转移两个电子，而这仅与两个ATP分子的水解相关联。在这里，我们测量了在稳态催化下，当铁蛋白簇处于[4Fe-4S]状态和处于[4Fe-4S]状态时，每次电子转移所消耗的ATP数量。结果发现，铁蛋白的两种氧化态在每次单电子转移事件中都是通过水解两个ATP来运作的。因此，无论其初始氧化还原状态如何，铁蛋白在将两个ATP水解的过程中，每次仅向钼铁蛋白转移一个电子。
