CHARACTERIZATION OF TRYPANOSOME ALTERNATIVE OXIDASE

非洲锥虫的生命周期以线粒体生物发生为特征，因为寄生虫从血流分化为亲周期的锥虫。血液生物体的特征是存在一种替代通常的细胞色素氧化酶的氧化酶。这种锥虫体交替氧化酶(TAO)是抗氰化物的，但被水杨基异羟肟酸100%抑制，水杨基异羟肟酸是植物和真菌中已知的交替氧化酶的抑制剂。与采采蝇中肠中发现的类似的亲周期类鞭毛虫，有75%的氰化物敏感的呼吸和25%的假敏感，表明在分化过程中TAO被保留。这种酶系统对血液寄生虫至关重要，因为它负责血流锥虫的有氧呼吸，它的抑制将损害寄生虫的生存机会。它在锥虫体内NADH的再氧化中起着至关重要的作用。我们最近克隆、测序并表达了布氏锥虫的TAO基因。这项建议的具体目的是：1.阐明TAO的蛋白质结构；2.阐明与TAO功能有关的关键性质；3.评估TAO对锥虫的重要性及其作为药物靶点的潜力。4.评价一种潜在的TAO抑制剂及其作为杀锥虫药物的有效性。这项研究的目的是在分子和生化水平上表征锥虫体内的TAO，以便为将该酶用作杀锥虫药物靶标的研究提供有价值的信息。

The life cycle of African trypanosomes is characterized by mitochondrial biogenesis as the parasites differentiate from the bloodstream to pro- cyclic trypomastigotes. The bloodstream organisms are characterized by the presence of an alternative oxidase to the usual cytochrome oxidase. This trypanosome alternative oxidase (TAO) is cyanide-resistant but inhibited 100% by salicylhydroxamic acid, a known inhibitor of alternative oxidases in plants and fungi. Pro-cyclic trypomastigotes similar to those found in the mid-gut of the tsetse fly, have 75% cyanide- sensitive respiration and 25% SHAM sensitive suggesting a retention of the TAO during differentiation. This enzyme system is critically important to the bloodstream parasite as it is responsible for the aerobic respiration of the bloodstream trypanosomes and its inhibition would damage the parasite's chances for survival. It plays an essential role in the re-oxidation of NADH in the trypanosome. We have recently cloned, sequenced, and expressed the TAO genes from Trypanosoma brucei brucei. The specific aims of this proposal are: 1. To elucidate the protein structure of TAO; 2. To elucidate key properties responsible for the function of the TAO; and 3. To evaluate how essential the TAO is to the trypanosome and its potential as a drug target. 4. To evaluate new compounds a potential inhibitors of TAO and their effectiveness as trypanocidal drugs. The goal of the proposed research is to characterize the TAO in trypanosomes at the molecular and biochemical levels so as to have information valuable for studies leading to the use of this enzyme as a target for trypanocidal drugs.