STRUCTURAL STUDIES OF REDOX STATES OF MAUG

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。列出的机构是 研究中心，而研究中心不一定是研究者所在的机构。 该合作项目的目标是从结构上表征MauG的不同氧化还原状态。MauG是一种非常不寻常的二血红素酶，其完成了在甲胺脱氢酶（MADH）中发现的新型氨基酸衍生的催化辅因子色氨酸甲萘醌（TTQ）的合成。TTQ是通过MADH β多肽链中两个Trp残基的翻译后修饰形成的，在此过程中，两个氧原子掺入betaTrp 57的吲哚环中，并在betaTrp 57和betaTrp 108的吲哚环之间形成共价键。MauG（preMADH）的天然底物是MADH的119 kDa蛋白质前体，具有单羟基化β Trp 57且无交联。MauG催化六电子氧化以完成TTQ生物合成。它可以在H2 O2依赖性或O2/还原当量依赖性反应中实现这一点。该酶已被证明形成一个前所未有的二血红素双铁（IV）物种，是催化能力。该中间体与Fe（V）电子等价，并且由Fe（IV）=O和位于第二血红素的Fe上的第二氧化等价物组成。我们已经解决了MauG与preMADH复合物的晶体结构。这已经达到了2.1英寸的分辨率，这些晶体可以支持催化周转形成TTQ而不会损失衍射。我们预计Fe（IV）=O形成的5配位P-血红素是TTQ的32 <$。6-坐标E-血红素位于P-血红素和TTQ之间的等距，并且具有非常不寻常的His/Tyr轴向配体构型，我们假设这是稳定Fe（IV）所需的。使用XAS在溶液中和晶体中，我们希望表征的分子环境，可以支持和稳定的不寻常的氧化还原物种的MauG，并使我们能够作出根本性的发现有关的氧活化的一个前所未有的二血红素双铁（IV）中间体。这项工作将是XAS对这种不寻常的酶的第一次研究。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The objective of this collaborative project is to structurally characterize different redox states of MauG. MauG is a highly unusual di-heme enzyme that completes the synthesis of the novel amino acid derived catalytic cofactor, tryptophan tryptophylquinone (TTQ) found in the enzyme methylamine dehydrogenase (MADH). TTQ is formed by post-translational modification of two Trp residues in the MADH beta polypeptide chain during which two atoms of oxygen are incorporated into the indole ring of betaTrp57 and a covalent bond is formed between the indole rings of betaTrp57 and betaTrp108. The natural substrate for MauG (preMADH) is a 119 kDa protein precursor of MADH with mono-hydroxylated betaTrp57 and no cross-link. MauG catalyzes a six-electron oxidation to complete TTQ biosynthesis. It can do this in a H2O2-dependent or O2/reducing equivalents-dependent reaction. The enzyme has been shown to form a unprecedented di-heme bis-Fe(IV) species that is catalytically competent. This intermediate is electronically equivalent to Fe(V), and is composed of an Fe(IV)=O with the second oxidizing equivalent residing on the Fe of the second heme. We have solved the crystal structure of MauG in complex with preMADH. This has been achieved to a resolution of 2.1 ¿, and these crystals can support catalytic turnover to form TTQ without loss of diffraction. The 5-coordinate P-heme that we expect to be the site of Fe(IV)=O formation is 32 ¿ from TTQ. The 6-coordinate E-heme lies equidistant between the P-heme and TTQ, and has a highly unusual His/Tyr axial ligand configuration that we hypothesize is required for stabilizing Fe(IV). Using XAS both in solution and in the crystal, we wish to characterize the molecular environments that can support and stabilize the unusual redox species of MauG, and enable us to make fundamental discoveries about oxygen activation by an unprecedented di-heme bis-Fe(IV) intermediate. This work would be the first XAS study of this unusual enzyme.