Thiol redox control in the single mitochondrion of trypanosomes

锥虫是几种热带疾病的病原体，具有不寻常的基于锥虫硫酮的硫醇氧化还原代谢。寄生虫特异性代谢物由亚精胺和两个谷胱甘肽分子合成，并通过黄素酶锥虫硫酮还原酶保持还原。参与生物合成的所有酶以及锥虫硫酮还原酶仅存在于胞质溶胶中。在此，锥虫硫酮通过直接巯基/二硫化物交换还原谷胱甘肽二硫化物，并为各种氧化还原蛋白质（如Tryparedoxin、硫氧还蛋白和谷氧还蛋白）提供还原当量。对于任何锥虫微生物，均未在任何亚细胞器中鉴定出低分子量硫醇。本项目的目的是阐明谷胱甘肽和/或锥虫硫酮是否发生在布氏锥虫的阶段依赖性调节的单细胞中，硫醇如何转移到细胞器中，以及当寄生虫受到氧化应激时，细胞溶质和线粒体硫醇池的氧化还原平衡如何受到影响。第二种方法涉及的生理作用，可能是必不可少的线粒体硫氧还蛋白型蛋白，没有任何对应物以外的锥虫。这些研究将包括重组蛋白的结构和功能分析以及条件敲除寄生虫的产生和表征。计划中的工作应该为我们提供对这些寄生虫单个线粒体的巯基代谢的全面了解，并解开新型硫氧还蛋白是否是低分子量巯基和线粒体巯基过氧化物酶之间缺失的环节。

Trypanosomatids, to which belong the causative agents of several tropical diseases, have an unusual trypanothione-based thiol redox metabolism. The parasite-specific metabolite is synthesized from spermidine and two molecules of glutathione and kept reduced by the flavoenzyme trypanothione reductase. All enzymes involved in the biosynthesis as well as trypanothione reductase exclusively occur in the cytosol. Here, trypanothione reduces glutathione disulfide by direct thiol/disulfide exchange and provides the reducing equivalents for various redox proteins such as tryparedoxin, thioredoxin and glutaredoxin. For none of the trypanosomatid organism, the low molecular mass thiol(s) in any subcellular organelle has/have been identified. Aim of this project is to elucidate if glutathione and/or trypanothione occur in the stage-dependent regulated single mitochondrion of Trypanosoma brucei, how the thiols are transferred into the organelle and how the redox balance of the cytosolic and mitochondrial thiol pools are affected when the parasites are oxidatively stressed. The second approach deals with the physiological role of a probably essential mitochondrial thioredoxin-type protein which does not have any counterpart outside the trypanosomatids. These studies will comprise the structural and functional analysis of the recombinant protein as well as the generation and characterization of conditional knockout parasites. The planned work should provide us with a comprehensive insight into the thiol metabolism of the singular mitochondrion of these parasites and unravel if the novel thioredoxin is the missing link between the low molecular mass thiols and the mitochondrial thiol peroxidases.