Effect of a Tridentate Ligand on the Structure, Electronic Structure, and Reactivity of the Copper(I) Nitrite Complex: Role of the Conserved Three-Histidine Ligand Environment of the Type-2 Copper Site in Copper-Containing Nitrite Reductases

It is postulated that the copper(I) nitrite complex is a key reaction intermediate of copper containing nitrite reductases (Cu-NiRs), which catalyze the reduction of nitrite to nitric oxide (NO) gas in bacterial denitrification. To investigate the structure–function relationship of Cu-NiR, we prepared five new copper(I) nitrite complexes with sterically hindered tris(4-imidazolyl)carbinols [Et-TIC = tris(1-methyl-2-ethyl-4-imidazolyl)carbinol and iPr-TIC = tris(1-methyl-2-isopropyl-4-imidazolyl)carbinol] or tris(1-pyrazolyl)methanes [Me-TPM = tris(3,5-dimethyl-1-pyrazolyl)methane; Et-TPM = tris(3,5-diethyl-1-pyrazolyl)methane; and iPr-TPM = tris(3,5-diisopropyl-1-pyrazolyl)methane]. The X-ray crystal structures of all of these copper(I) nitrite complexes were mononuclear η1-N-bound nitrite complexes with a distorted tetrahedral geometry. The electronic structures of the complexes were investigated by absorption, magnetic circular dichroism (MCD), NMR, and vibrational spectroscopy. All of these complexes are good functional models of Cu-NiR that form NO and copper(II) acetate complexes well from reactions with acetic acid under anaerobic conditions. A comparison of the reactivity of these complexes, including previously reported (iPr-TACN)Cu(NO2) [iPr-TACN = 1,4,7-triisopropyl-1,4,7-triazacyclononane], clearly shows the drastic effects of the tridentate ligand on Cu-NiR activity. The copper(I) nitrite complex with the Et-TIC ligand, which is similar to the highly conserved three-histidine ((His)3) ligand environment in the catalytic site of Cu-NiR, had the highest Cu-NiR activity. This result suggests that the (His)3 ligand environment is essential for acceleration of the Cu-NiR reaction. The highest Cu-NiR activity for the Et-TIC complex can be explained by the structural and spectroscopic characterizations and the molecular orbital calculations presented in this paper. Based on these results, the functional role of the (His)3 ligand environment in Cu-NiR is discussed.

有研究提出，亚硝酸铜(I)配合物是含铜亚硝酸还原酶（Cu-NiRs）的关键反应中间体，该类酶可在细菌反硝化过程中催化亚硝酸根还原为一氧化氮（NO）气体。为探究含铜亚硝酸还原酶的构效关系，我们合成了5种新型位阻型亚硝酸铜(I)配合物，其配体分别为位阻型三(4-咪唑基)甲醇类配体[三(1-甲基-2-乙基-4-咪唑基)甲醇（Et-TIC）、三(1-甲基-2-异丙基-4-咪唑基)甲醇（iPr-TIC）]或三(1-吡唑基)甲烷类配体[三(3,5-二甲基-1-吡唑基)甲烷（Me-TPM）、三(3,5-二乙基-1-吡唑基)甲烷（Et-TPM）、三(3,5-二异丙基-1-吡唑基)甲烷（iPr-TPM）]。所有上述亚硝酸铜(I)配合物的X射线晶体结构均为单核η1-N配位的亚硝酸根配合物，具有畸变四面体几何构型。我们通过吸收光谱、磁圆二色谱（MCD）、核磁共振波谱（NMR）以及振动光谱对配合物的电子结构进行了表征。在厌氧条件下，此类配合物均可与乙酸反应生成一氧化氮与乙酸铜(II)配合物，是一类性能优异的含铜亚硝酸还原酶功能模型。通过对比本研究中配合物与已有报道的(1,4,7-三异丙基-1,4,7-三氮杂环壬烷)亚硝酸铜(I)配合物[(iPr-TACN)Cu(NO2)]的反应活性，可明确三齿配体对含铜亚硝酸还原酶活性的显著调控作用。其中，搭载Et-TIC配体的亚硝酸铜(I)配合物，其配位环境与含铜亚硝酸还原酶催化位点中高度保守的三组胺酸（(His)3）配位环境极为相似，展现出最高的含铜亚硝酸还原酶活性。该结果表明，三组胺酸配位环境对于加速含铜亚硝酸还原酶的催化反应至关重要。本研究通过结构表征、光谱分析以及分子轨道计算，阐明了Et-TIC配合物具备最高催化活性的内在机制。基于上述结果，我们进一步讨论了三组胺酸配位环境在含铜亚硝酸还原酶中的功能作用。