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，寄生虫从血流向前循环锥虫分化。血流生物体的特点是存在一种替代通常细胞色素氧化酶的氧化酶。这种锥虫替代性氧化酶（TAO）是抗氰化物的，但被水杨基羟肟酸100%抑制，水杨基羟肟酸是植物和真菌中已知的替代性氧化酶抑制剂。与在采采蝇中肠中发现的类似，前环锥乳线虫具有75%的氰化物敏感呼吸和25%的SHAM敏感呼吸，这表明在分化过程中TAO保留。这种酶系统对血流寄生虫至关重要，因为它负责血流锥虫的有氧呼吸，抑制它会损害寄生虫的生存机会。它在锥虫体内NADH的再氧化中起重要作用。我们最近克隆、测序和表达了来自布鲁氏锥虫的TAO基因。本建议的具体目的是：1。阐明TAO的蛋白结构；2. 阐明TAO功能的关键属性；和3。评估TAO对锥虫的重要性及其作为药物靶点的潜力。4. 评价潜在的TAO抑制剂新化合物及其作为锥虫药物的有效性。本研究的目的是在分子和生化水平上对锥虫体内的TAO进行表征，从而为利用该酶作为锥虫药物靶点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.