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Structural study of the activation mechanism of dihydrogen at the Ni-Fe active site of [NiFe] hydrogenase

[NiFe]氢化酶Ni-Fe活性位点氢气激活机制的结构研究

基本信息

批准号：
14380317
负责人：
HIGUCHI Yoshiki
金额：
$ 7.68万
依托单位：
University of Hyogo (2004)Himeji Institute of Technology (2002-2003)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

项目摘要

Hydrogenases catalyze the reversible oxidation of molecular hydrogen and play a key role in hydrogen metabolism in various bacteria. Dihydrogen, the substrate and product of hydrogenases, is a good candidate for fuel in the future, since fossil fuels are a limited resource and considered to cause environmental disruption. The reaction mechanism of dihydrogen production by hydrogenases is potentially useful for development of new chemical engineering processes for hydrogen fuels, whereas that of dihydrogen consumption is potentially applicable for new types of fuel cells. While various lines of evidence indicated that the extrinsic CO interacts with the Ni atom of [NiFe]hydrogenase, they were not entirely conclusive, given the hetero-binulcear nature of the active site and the intimate communication between the two metals. In this study, we present the first direct evidence of CO coordination to the Ni atom of D.V.Miyazaki [NiFe]hydrogenase by X-ray crystallography.The carbon monoxide c … More omplex of [NiFe] hydrogenase from Desulfovibrio vulgaris Miyazaki F has been characterized by X-ray crystallography and absorption and resonance Raman spectroscopy. Nine crystal structures of the [NiFe] hydrogenase in the CO-bound and CO-liberated forms were determined at 1.2-1.4 Å resolution. The exogenously added CO was assigned to be bound to the Ni atom at the Ni-Fe active site. Distinct changes were observed in the electron density distribution of the Ni and Sγ(Cys546) atoms between the CO-bound and CO-liberated structures for all the crystals tested. The novel structural features found near the Ni and Sγ(Cys546) atoms suggest that these two atoms at the Ni-Fe active site play a role during the initial H2-binding process.[NiFe] hydrogenase has two different oxidized states, Ni-A (unready) and Ni-B (ready). We have succeeded in converting Ni-B to Ni-A with the use of Na_2S and O_2, and determination of the high resolution crystal structures of the both states. Ni-B possesses a monatomic non-protein bridging ligand at the Ni-Fe active site, whereas Ni-A has a diatomic species. The terminal atom of the bridging species of Ni-A occupies a similar position as C of the exogenous CO in the CO complex (inhibited state). The common features of the enzyme structures at the resting (Ni-A) and inhibited (CO complex) states are proposed. These findings provide useful information on the design of new systems of biomimetic dihydrogen production and fuel cell devices. Less

氢化酶催化分子氢的可逆氧化，在各种细菌的氢代谢中起关键作用。氢酶的底物和产物二氢是未来燃料的良好候选者，因为化石燃料是有限的资源并且被认为会引起环境破坏。氢化酶产生氢的反应机理对开发新的氢燃料化学工程过程具有潜在的意义，而消耗氢的反应机理对新型燃料电池具有潜在的应用价值。虽然各种证据表明，外来CO与[NiFe]氢化酶的Ni原子相互作用，但考虑到活性位点的异双核性质和两种金属之间的密切联系，它们并不完全是决定性的。本研究首次用X射线晶体学方法直接证明了D. V.宫崎[NiFe]氢化酶中的Ni原子与CO配位，并对CO的配位机理进行了初步探讨。 ...更多信息用X射线晶体学、吸收光谱和共振拉曼光谱对脱硫弧菌（Desulfovibrio vulgaris）宫崎F的[NiFe]氢化酶复合物进行了表征。在1.2-1.4 μ m分辨率下测定了9种CO结合和CO释放形式的[NiFe]氢化酶的晶体结构。外加CO与Ni原子结合在Ni-Fe活性位上。对于所有测试的晶体，在CO结合和CO释放结构之间观察到Ni和Sγ（Cys 546）原子的电子密度分布的明显变化。在Ni和Sγ（Cys 546）原子附近发现的新结构特征表明，Ni-Fe活性中心的这两个原子在初始H2结合过程中起作用。[NiFe]氢化酶有两种不同的氧化态，Ni-A（未准备好）和Ni-B（准备好）。我们用Na_2S和O_2成功地将Ni-B转变为Ni-A，并测定了这两种状态的高分辨晶体结构。Ni-B在Ni-Fe活性位点具有单原子非蛋白质桥连配体，而Ni-A具有双原子物种。桥物种的Ni-A的末端原子在CO络合物中占据与外源CO的C类似的位置（抑制状态）。的酶结构在休息（镍A）和抑制（CO复合物）状态的共同特点提出。这些发现为仿生制氢新系统和燃料电池装置的设计提供了有用的信息。少