权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

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

氢化酶催化氢分子的可逆氧化，在各种细菌的氢代谢中发挥关键作用。氢气是氢化酶的底物和产物，是未来燃料的良好候选者，因为化石燃料是有限的资源，并且被认为会造成环境破坏。氢化酶产生氢气的反应机制可能有助于开发氢燃料的新化学工程工艺，而氢气消耗的反应机制则可能适用于新型燃料电池。虽然各种证据表明外在 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以及两种金属之间的密切联系，这些证据并不完全是结论性的。在这项研究中，我们通过 X 射线晶体学首次提供了 CO 与 D.V.Miyazaki [NiFe] 氢化酶的 Ni 原子配位的直接证据。来自普通脱硫弧菌 Miyazaki F 的 [NiFe] 氢化酶的一氧化碳复合物已通过 X 射线晶体学以及吸收和共振拉曼光谱进行了表征。 CO 结合型和 CO 释放型的 [NiFe] 氢化酶的九种晶体结构以 1.2-1.4 Å 分辨率测定。外源添加的 CO 被认为与 Ni-Fe 活性位点的 Ni 原子结合。对于所有测试的晶体，在 CO 结合结构和 CO 释放结构之间的 Ni 和 Sγ(Cys546) 原子的电子密度分布观察到明显的变化。在 Ni 和 Sγ(Cys546) 原子附近发现的新结构特征表明，Ni-Fe 活性位点的这两个原子在最初的 H2 结合过程中发挥了作用。[NiFe]氢化酶具有两种不同的氧化态：Ni-A（未就绪）和 Ni-B（就绪）。我们利用Na_2S和O_2成功地将Ni-B转化为Ni-A，并测定了两种状态的高分辨率晶体结构。 Ni-B 在 Ni-Fe 活性位点具有单原子非蛋白质桥配体，而 Ni-A 具有双原子种类。 Ni-A 桥连物种的末端原子与 CO 复合物中外源 CO 的 C 占据相似的位置（抑制状态）。提出了静息（Ni-A）和抑制（CO 复合物）状态下酶结构的共同特征。这些发现为仿生氢气生产和燃料电池装置的新系统的设计提供了有用的信息。较少的