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Synthesis and reactivity of synthetic analogs for nickel redox enzymes: Superoxide dismutase and acetyl coenzyme A synthase.

机译：镍氧化还原酶的合成类似物的合成和反应性：超氧化物歧化酶和乙酰​​辅酶A合酶。

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摘要

Synthetic model complexes of nickel-containing enzyme active sites are important tools for understanding the structure-function properties of the active sites as well as for investigating the enzyme mechanisms. The synthesis and reactivity of structural mimics and catalytic-intermediate analogs can provide insight into how the diverse chemistry of nickel-containing enzymes occurs. Reported herein are two synthetic models of redox active nickel-dependant metalloproteins, nickel-containing superoxide dismutase and acetyl coenzyme A synthase.;Nickel-containing superoxide dismutase, Ni SOD, contains a mononuclear nickel active site found at the N-terminus with active site ligands found within the first six residues of the protein sequence.1 The active site sequence is unstructured in the absence of Ni2+ providing a novel target for synthetic modeling. A Ni SOD model system was designed utilizing short peptides as ligands based on the native active site amino acid sequence, NH2-His-Cys-Gly-Gly-Pro-Cys-COOH. The system is used to investigate the minimum construct necessary to establish the native peptide fold and metal coordination.;The [Ni(HCGGPC)]- complex was determined to be monomeric by mass spectrometry and optical spectral titrations with K d = 4.7 +/- 0.4 x 10-12 M. One- and two-dimensional NMR spectral data suggests the nickel coordination sphere is the same as found in the enzyme. Metal selectivity of the peptide was measured comparing Ni 2+, Zn2+, and Co2+. Ni2+ is preferred two orders of magnitude over Zn2+ and three orders of magnitude over Co2+. The reactivity of the model complex with O2·- is similar to that observed for the enzyme.1 Reaction of the complex with substoichiometric KO2 results in a Ni3+ species with a similar EPR spectrum as that observed in the enzyme. The superoxide dismutase activity of the nickel complex was measured to be, IC50=191 +/- 9 muM much reduced compared to the native enzyme. The decreased activity is due to the product-catalyzed decomposition of the complex suggesting substrate channeling and the active site pocket of the enzyme provide essential oxidative protection of the active site thiols.;Acetyl coenzyme A synthase, ACS, catalyzes the production of acetyl coenzyme A from one-carbon units, CO and CH3. A recently proposed ACS catalytic mechanism involves a zero valent nickel as the complex to accept a methyl group from methylcobalt species during catalytic turnover.2 Synthetic analog studies were designed to probe the feasibility of Ni(0) as the methyl acceptor. Reaction of [triphos]Ni(PPh3) (triphos = bis(2-diphenylphosphinoethyl)phenyl phosphine) with CH3Co(dmgBF2)2py proceeds quantitatively yielding [Ni(triphos)(Me)]+ and Co(I). 3 Kinetic data of methyl transfer and the rate of other alkyls (Et, iPr) suggests that the mechanism of alkyl transfer proceeds via an SN2 pathway similar to the ACS enzyme. The nickel-alkyls complexes possessing beta-hydrogen undergo beta-hydrogen elimination forming a nickel-hydride complex characterized by NMR, FT-IR spectroscopies and X-ray crystallography. This system is also used to model another intermediate along the catalytic pathway, the Ni-acyl complex, to help determined the importance of the order of substrate binding to the ACS active site during catalysis.
机译：含镍酶活性位点的合成模型复合物是理解活性位点的结构功能特性以及研究酶机理的重要工具。结构模拟物和催化中间体类似物的合成和反应性可以提供洞察力，以了解含镍酶的各种化学反应是如何发生的。本文报道了氧化还原活性镍依赖性金属蛋白的两种合成模型，即含镍超氧化物歧化酶和乙酰​​辅酶A合酶;含镍超氧化物歧化酶Ni SOD包含在N端具有活性位点的单核镍活性位点在蛋白质序列的前六个残基中发现了一些配体。1在没有Ni2 +的情况下，活性位点序列是无结构的，为合成建模提供了新的靶标。基于天然活性位点氨基酸序列NH2-His-Cys-Gly-Gly-Pro-Cys-COOH，使用短肽作为配体设计了Ni SOD模型系统。该系统用于研究建立天然肽折叠和金属配位所需的最小构建体。; [Ni（HCGGPC）]-络合物经质谱和光谱滴定法确定为单体，K d = 4.7 +/- 0.4 x 10-12M。一维和二维NMR光谱数据表明镍配位球与酶中的相同。比较Ni 2 +，Zn2 +和Co2 +来测量肽的金属选择性。 Ni 2+优选比Zn 2+高两个数量级，并且比Co 2+高三个数量级。模型络合物与O2·-的反应性与酶类似。1络合物与亚化学计量的KO2的反应产生的Ni3 +物种的EPR谱与酶中观察到的相似。与天然酶相比，镍配合物的超氧化物歧化酶活性测得IC50 = 191 +/- 9μM。活性降低是由于复合物的产物催化分解，表明底物通道化，并且该酶的活性位点口袋为活性位点硫醇提供了重要的氧化保护。乙酰辅酶A合酶ACS催化乙酰辅酶A的产生由一碳单元CO和CH3最近提出的ACS催化机理涉及以零价镍作为络合物在催化周转期间接受甲基钴物种的甲基。2设计了合成模拟研究以探讨Ni（0）作为甲基受体的可行性。 [triphos] Ni（PPh3）（triphos =双（2-二苯基膦基乙基）苯基膦）与CH3Co（dmgBF2）2py的反应定量产生[Ni（triphos）（Me）] +和Co（I）。 3甲基转移的动力学数据和其他烷基的比率（Et，iPr）表明，烷基转移的机制是通过类似于ACS酶的SN2途径进行的。具有β-氢的镍-烷基络合物经过β-氢消除，形成了以NMR，FT-IR光谱和X射线晶体学为特征的氢化镍络合物。该系统还用于模拟催化途径中的另一种中间体Ni-酰基配合物，以帮助确定催化过程中底物与ACS活性位点结合顺序的重要性。

著录项

  • 作者

    O'Hagan, Molly J.;

  • 作者单位

    University of Delaware.;

  • 授予单位 University of Delaware.;
  • 学科 Chemistry Biochemistry.;Chemistry Inorganic.
  • 学位 D.A.
  • 年度 2010
  • 页码 188 p.
  • 总页数 188
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

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