School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Australia.
Acc Chem Res. 2012 Sep 18;45(9):1593-603. doi: 10.1021/ar300067g. Epub 2012 Jun 14.
Binuclear metallohydrolases are a large family of enzymes that require two closely spaced transition metal ions to carry out a plethora of hydrolytic reactions. Representatives include purple acid phosphatases (PAPs), enzymes that play a role in bone metabolism and are the only member of this family with a heterovalent binuclear center in the active form (Fe(3+)-M(2+), M = Fe, Zn, Mn). Other members of this family are urease, which contains a di-Ni(2+) center and catalyzes the breakdown of urea, arginase, which contains a di-Mn(2+) center and catalyzes the final step in the urea cycle, and the metallo-β-lactamases, which contain a di-Zn(2+) center and are virulence factors contributing to the spread of antibiotic-resistant pathogens. Binuclear metallohydrolases catalyze numerous vital reactions and are potential targets of drugs against a wide variety of human disorders including osteoporosis, various cancers, antibiotic resistance, and erectile dysfunctions. These enzymes also tend to catalyze more than one reaction. An example is an organophosphate (OP)-degrading enzyme from Enterobacter aerogenes (GpdQ). Although GpdQ is part of a pathway that is used by bacteria to degrade glycerolphosphoesters, it hydrolyzes a variety of other phosphodiesters and displays low levels of activity against phosphomono- and triesters. Such a promiscuous nature may have assisted the apparent recent evolution of some binuclear metallohydrolases to deal with situations created by human intervention such as OP pesticides in the environment. OP pesticides were first used approximately 70 years ago, and therefore the enzymes that bacteria use to degrade them must have evolved very quickly on the evolutionary time scale. The promiscuous nature of enzymes such as GpdQ makes them ideal candidates for the application of directed evolution to produce new enzymes that can be used in bioremediation and against chemical warfare. In this Account, we review the mechanisms employed by binuclear metallohydrolases and use PAP, the OP-degrading enzyme from Agrobacterium radiobacter (OPDA), and GpdQ as representative systems because they illustrate both the diversity and similarity of the reactions catalyzed by this family of enzymes. The majority of binuclear metallohydrolases utilize metal ion-activated water molecules as nucleophiles to initiate hydrolysis, while some, such as alkaline phosphatase, employ an intrinsic polar amino acid. Here we only focus on catalytic strategies applied by the former group.
双核金属水解酶是一大类酶，需要两个紧密间隔的过渡金属离子才能进行多种水解反应。代表性的酶包括紫色酸性磷酸酶 (PAP)，这种酶在骨代谢中发挥作用，是该家族中唯一在活性形式下具有杂价双核中心的成员（Fe(3+)-M(2+)，M = Fe、Zn、Mn）。该家族的其他成员包括脲酶，它含有一个双 Ni(2+)中心，催化尿素的分解；精氨酸酶，它含有一个双 Mn(2+)中心，催化尿素循环的最后一步；以及金属β-内酰胺酶，它含有一个双 Zn(2+)中心，是导致抗生素耐药病原体传播的毒力因子。双核金属水解酶催化许多重要的反应，是针对包括骨质疏松症、各种癌症、抗生素耐药性和勃起功能障碍等多种人类疾病的药物的潜在靶点。这些酶也往往催化不止一种反应。一个例子是来自 Aerogenes 肠杆菌的有机磷 (OP) 降解酶（GpdQ）。尽管 GpdQ 是细菌用于降解甘油磷酸酯途径的一部分，但它也能水解多种其他磷酸二酯，并对磷酸单酯和三酯显示出低水平的活性。这种混杂的性质可能有助于一些双核金属水解酶最近的进化，以应对人类干预（如环境中的 OP 农药）所造成的情况。OP 农药于大约 70 年前首次使用，因此细菌用于降解它们的酶必须在进化时间尺度上非常迅速地进化。像 GpdQ 这样的酶的混杂性质使它们成为定向进化的理想候选者，以产生可用于生物修复和对抗化学战的新酶。在本报告中，我们综述了双核金属水解酶所采用的机制，并以 PAP（来自 Agrobacterium radiobacter 的 OP 降解酶）和 GpdQ 为例，因为它们说明了该家族酶催化的反应的多样性和相似性。大多数双核金属水解酶利用金属离子激活的水分子作为亲核试剂来启动水解，而有些酶，如碱性磷酸酶，则利用内在极性氨基酸。在这里，我们只关注前一组酶所应用的催化策略。
