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首页> 外文期刊>Biochemistry >Electron/proton coupling in bacterial nitric oxide reductase during reduction of oxygen
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Electron/proton coupling in bacterial nitric oxide reductase during reduction of oxygen

机译：氧还原过程中细菌一氧化氮还原酶中电子/质子的耦合

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The respiratory nitric oxide reductase (NOR) from Paracoccus denitrificans catalyzes the two-electron reduction of NO to N2O (2NO + 2H(+) + 2e(-) -> N2O + H2O), which is an obligatory step in the sequential reduction of nitrate to dinitrogen known as denitrification. NOR has four redox-active cofactors, namely, two low-spin hemes c and b, one high-spin heme b(3), and a non-heme iron FeB, and belongs to same superfamily as the oxygen-reducing heme-copper oxidases. NOR can also use oxygen as an electron acceptor; this catalytic activity was investigated in this study. We show that the product in the steady-state reduction of oxygen is water. A single turnover of the fully reduced NOR with oxygen was initiated using the flow-flash technique, and the progress of the reaction monitored by time-resolved optical absorption spectroscopy. Two major phases with time constants of 40 mu s and 25 ms (pH 7.5, 1 mm O-2) were observed. The rate constant for the faster process was dependent on the O-2 concentration and is assigned to O-2 binding to heme b(3) at a bimolecular rate constant of 2 x 10(7) M-1 s(-1). The second phase (tau = 25 ms) involves oxidation of the low-spin hemes b and c, and is coupled to the uptake of protons from the bulk solution. The rate constant for this phase shows a pH dependence consistent with rate limitation by proton transfer from an internal group with a pK(a) = 6.6. This group is presumably an amino acid residue that is crucial for proton transfer to the catalytic site also during NO reduction.
机译：脱氮副球菌的呼吸型一氧化氮还原酶（NOR）催化将NO的两电子还原成N2O（2NO + 2H（+）+ 2e（-）-> N2O + H2O），这是依次还原N2O的必不可少的步骤。硝酸盐将二氮称为反硝化作用。 NOR具有四个氧化还原活性辅助因子，即两个低旋血红素c和b，一个高旋血红素b（3）和一个非血红素铁FeB，与降氧血红素铜属于同一超家族氧化酶。 NOR也可以使用氧作为电子受体。在这项研究中研究了这种催化活性。我们表明，稳态还原的产物是水。使用流闪技术开始完全还原的含氧NOR的单周转，并通过时间分辨光吸收光谱法监测反应进程。观察到两个主要相，时间常数分别为40μs和25 ms（pH 7.5，1 mm O-2）。更快过程的速率常数取决于O-2浓度，并以2 x 10（7）M-1 s（-1）的双分子速率常数分配给O-2与血红素b（3）结合。第二阶段（tau = 25 ms）涉及低自旋血红素b和c的氧化，并与从整体溶液中吸收质子有关。此阶段的速率常数显示出与pH的依赖性，该依赖性与pK（a）= 6.6的内部基团通过质子转移引起的速率限制相一致。该基团大概是氨基酸残基，对于NO还原过程中质子转移到催化位点也是至关重要的。

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