DETERMINATION OF THE METAL-ION SEPARATION AND ENERGIES OF THE 3 LOWEST ELECTRONIC STATES OF DIMANGANESE(II,II) COMPLEXES AND ENZYMES - CATALASE AND LIVER ARGINASE

DETERMINATION OF THE METAL-ION SEPARATION AND ENERGIES OF THE 3 LOWEST ELECTRONIC STATES OF DIMANGANESE(II,II) COMPLEXES AND ENZYMES - CATALASE AND LIVER ARGINASE
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DOI：
10.1021/bi00006a023
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发表时间：
1995-02-14
期刊：
BIOCHEMISTRY
影响因子：
2.9
通讯作者：
DISMUKES, GC
DISMUKES, GC
中科院分区：
生物学3区
文献类型：
--
作者：
KHANGULOV, SV;PESSIKI, PJ;DISMUKES, GC

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来自嗜热栖热菌的二锰（II，II）过氧化氢酶MnCat（II，II）、来自大鼠肝脏的过氧化氢酶Arg（II，II）和几种二锰（II，II）化合物LMn（2）XY（2）（它们是功能性过氧化氢酶模拟物）都在其催化位点具有一对偶联的Mn（II）离子。对于每一个这些，我们已经测量了EPR光谱的相对能量分离的三个最低的电子状态（单重态，三重态，五重态），描述了一个通用的方法提取这些国家的多组分分析的个人光谱，并确定了锰锰分离。三重态-单重态和五重态-单重态能隙通过将EPR强度的温度依赖性拟合到通过各向同性海森堡自旋交换耦合的一对Mn（II）离子的Boltzmann表达式（-2JS（1）S（2））来很好地建模。这种依赖性表明，对于Arg（II，II）（+硼酸盐）和MnCat（II，II）（磷酸盐），抗磁性基态分别具有Δ E（10）（cm（-1））= 12 J = 4和11.2cm（-1）。这种大的差异反映了桥连配体的差异，或者可能反映了酶中Mn（II）离子的配体场较弱（电离势较大）。在正丁醇/CH_2Cl_2中，[LMn（2）（CH_3CO_2）]（Cl_4）（2）（1），[LMn（2）（CH_3CO_2）（3）]（2）和[LMn（2）Cl（3）]（3）（HL = N，N，N ′，N ′-四（2-亚甲基苯并咪唑）-1，3-二氨基丙-2-醇）的三重态-单重态能隙为23-24 cm（-1）。海森堡交换相互作用常数的比较超过30个二锰（II，II）配合物表明可能的桥接结构（μ-OH）（μ-羧酸）（1-2）MnCat（II，II），而3倍较弱的耦合在Arg（II，II）表明μ-水代替μ-氢氧化物。提取了三重态和五重态电子态的EPR谱，发现它们表现出零场分裂（ZFS），并分辨了Mn-55超精细分裂，表明存在自旋耦合的Mn-2（II，II）物种。Mn（II）离子间的磁偶极-偶极相互作用是主要的磁相互作用。晶体学确定的Mn-Mn的距离和五重态（D-2）之间观察到的线性相关性的五个二锰对，这两个数据集是可用的。使用该相关性，预测Arg（II，II）中的Mn-Mn距离对于天然酶（多种形式）为3.36-3.57埃，并且对于MnCat（II，II）（磷酸盐）为3.59埃。将抑制剂硼酸盐添加到Arg（II，II）中简化了结晶，表明转化为单一物质，平均Mn-Mn分离为3.50埃。双核配合物中的第二个金属离子拥有一个共享的桥接配体已被证明削弱的μ-配体场电位的强度，监测的单离子的强度。减弱的μ-配体场电位可以发挥作用，例如，在促进质子从锰结合的水分子中离子化的酶，与所得的氢氧化物配体是水解底物所需的亲核试剂。
The dimanganese(II,II) catalase from Thermus thermophilus, MnCat(II,II), arginase from rat liver, Arg(II,II), and several dimanganese(II,II) compounds, LMn(2)XY(2), which are functional catalase mimics, all possess a pair of coupled Mn(II) ions in their catalytic sites. For each of these, we have measured by EPR spectroscopy the relative energies separating the three lowest electronic states (singlet, triplet, and quintet), described a general method for extracting the individual spectra for these states by multicomponent analysis, and determined the Mn-Mn separation. The triplet-singlet and quintet-singlet energy gaps were modeled well by fitting the temperature dependence of the EPR intensities to a Boltzmann expression for a pair of Mn(II) ions coupled by isotropic Heisenberg spin exchange (-2JS(1)S(2)). This dependence indicates diamagnetic ground states with Delta E(10) (cm(-1)) = \2J\ = 4 and 11.2 cm(-1) for Arg(II,II)(+borate) and MnCat(II,II)(phosphate), respectively. This large difference in \2J\ reflects either a difference in the bridging ligands or, possibly, a weaker ligand field (larger ionization potential) for the Mn(II) ions in arginase. In n-butanol/CH2Cl2 the triplet-singlet energy gaps for [LMn(2)(CH3CO2)](ClO4)(2) (1), [LMn(2)(CH3CO2)(3)] (2), and [LMn(2)Cl(3)] (3), where HL = N,N,N',N'-tetrakis(2-methylenebenzimidazole)-1,3-diaminopropan-2-ol, are 23-24 cm(-1). Comparison of the Heisenberg exchange interaction constants for more than 30 dimanganese(II,II) complexes suggests a possible bridging structure of (mu-OH)(mu-carboxylate)(1-2) for MnCat(II,II), while the 3-fold weaker coupling in Arg(II,II) suggests mu-aqua in place of mu-hydroxide. EPR spectra of both the triplet and quintet electronic states were extracted and found to exhibit zero-field splittings (ZFS) and resolved Mn-55 hyperfine splittings indicating spin-coupled Mn-2(II,II) species. The major ZFS interaction could be attributed to the magnetic dipole-dipole interaction between the Mn(II) ions. A linear correlation is observed between the crystallographically determined Mn-Mn distance and the ZFS of the quintet state (D-2) for five dimanganese pairs for which both data sets are available. Using this correlation, the Mn-Mn distance in Arg(II,II) is predicted to be 3.36-3.57 Angstrom for the native enzyme (multiple forms) and 3.59 Angstrom for MnCat(II,II)(phosphate). Addition of the inhibitor borate to Arg(II,II) simplifies the ZFS, indicative of conversion to a single species with mean Mn-Mn separation of 3.50 Angstrom. The second metal ion in dinuclear complexes possessing a shared bridging ligand has been shown to attenuate the strength of the mu-ligand field potential, as monitored by the strength of the single ion ZFS. A weakened mu-ligand field potential may play a role, for example, in promoting ionization of a proton from a manganese-bound water molecule in arginase, with the resulting hydroxide ligand being the nucleophile needed for hydrolysis of substrate.