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促周期锥鞭毛体。血流生物体的特征在于存在通常的细胞色素氧化酶的替代氧化酶。这种锥虫交替氧化酶（TAO）是抗氰化物的，但抑制100%的水杨基异羟肟酸，一种已知的植物和真菌中的交替氧化酶的抑制剂。类似于采采蝇中肠中发现的那些的前环状锥鞭毛体具有75%的氰化物敏感性呼吸和25%的SHAM敏感性，表明在分化期间TAO的保留。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，并且其抑制将损害寄生虫的存活机会。它在锥虫中NADH的再氧化中起重要作用。我们最近从布氏锥虫中克隆、测序并表达了TAO基因。该提案的具体目标是：1.阐明TAO的蛋白质结构; 2.阐明TAO功能的关键特性; 3.评价TAO对锥虫的重要性及其作为药物靶点的潜力。4.评价新化合物作为TAO抑制剂的潜力及其作为杀锥虫药物的有效性.拟议研究的目标是在分子和生物化学水平上表征锥虫中的TAO，以便为使用这种酶作为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.