4-Hydroxyphenylpyruvate Dioxygenase and Tyrosinemia

说明(申请人提供)：铁依赖的非血红素加氧酶是 本研究的具体着力点。这些酶催化重要的代谢和 分解代谢反应贯穿整个哺乳动物的新陈代谢。最初的意图是 通过以下方式发展对这些酶的化学的基本了解 鉴定过程中产生的瞬时氧中间体 催化作用。我们的长期目标是找到证据来证明 特定催化步骤的过渡态，以使稳定的过渡 状态模拟物可以被开发为抑制治疗药物来控制 通过关键的新陈代谢途径进行流动。最初，这种酶 选择了4-羟基苯丙酮酸双加氧酶(HPPD)作为研究对象。 这种酶体现了其他依赖FeII的酶的许多催化功能 芳香族加氧酶、氧化脱羧酶等非血红素双加氧酶 和取代基迁移。此外，它是为数不多的阿尔法酮之一 晶体结构已知的依赖酸的加氧酶。此外， 它已经成为缓解新陈代谢能力的典范 通过选择性的酶抑制而引起的疾病。 1型酪氨酸血症是由活性延胡索乙酸乙酯酶缺乏引起的 催化分解代谢途径中最后一步的酶 酪氨酸。在缺乏治疗的情况下，这种疾病往往一开始就是致命的 这是生命的一年，而且总是在前二十年完成之前。它是 然而，众所周知，HPPD的抑制催化了第二步 在大多数情况下，这种途径是治疗1型酪氨酸血症的有效方法。 这项研究的部分目的是，这种抑制的机制是 在酶的催化事件的背景下理解。实验性的 方法是将稳态和预稳态分析与 选择性使用底物类似物、同位素标记和诱变以及 在速率常数的基础上测试机械假说的结晶学 调制和结构。

DESCRIPTION (provided by applicant): Iron-dependent non-heme oxygenases are the specific focus of this research. These enzymes catalyze vital metabolic and catabolic reactions throughout mammalian metabolism. The initial intent is to develop a fundamental understanding of the chemistry of these enzymes by the identification of the transient oxygen intermediates that are generated during catalysis. The long-term objective is to find evidence for the geometry of transition states for specific catalytic steps such that stable transition state mimics can be developed as inhibitory therapeutic agents to control the flux through key metabolic pathways. Initially, the enzyme 4-hydroxyphenylpyruvate dioxygenase (HPPD) has been selected for investigation. This enzyme exemplifies many of the catalytic functions of other FeII-dependent non-heme dioxygenases such as aromatic oxygenation, oxidative decarboxylation and substituent migration. Furthermore, it is one of the few alpha-keto acid-dependent oxygenases for which the crystal structure is known. Moreover, it has become a paradigm example of the ability to alleviate metabolic disorders through selective enzymatic inhibition. Type 1 Tyrosinemia, is caused by a deficiency of active fumarylacetoacetase, an enzyme that catalyzes the final step in the pathway for the catabolism of tyrosine. In the absence of treatment this disease is often fatal in the first year of life and always prior the completion of the first two decades. It is known, however, that the inhibition of HPPD which catalyzes the second step of this pathway is, in most cases, an effective treatment for type 1 Tyrosinemia. Part of the intent of this research is that the mechanism of this inhibition be understood in the context of catalytic events of the enzyme. The experimental approach will be to combine steady state and pre-steady state analyses with the selective use of substrate analogs, isotopic labels and mutagenesis and crystallography to test mechanistic hypotheses on the basis of rate constant modulation and structure.