RESULTS: We report unprecedented modification of TIM in hydrogenosomes of the human parasite Trichomonas vaginalis (TvTIM). We show that the import of the presequence-containing protein into the hydrogenosomal matrix is mediated by the hybrid TIM22-TIM23 complex that includes three highly divergent core components, TvTim22, TvTim23, and TvTim17-like proteins. The hybrid character of the TvTIM is underlined by the presence of both TvTim22 and TvTim17/23, association with small Tim chaperones (Tim9-10), which in mitochondria are known to facilitate the transfer of substrates to the TIM22 complex, and the coupling with TIM23-specific ATP-dependent presequence translocase-associated motor (PAM). Interactome reconstruction based on co-immunoprecipitation (coIP) and mass spectrometry revealed that hybrid TvTIM is formed with the compositional variations of paralogs. Single-particle electron microscopy for the 132-kDa purified TvTIM revealed the presence of a single ring of small Tims complex, while mitochondrial TIM22 complex bears twin small Tims hexamer. TvTIM is currently the only TIM visualized outside of Opisthokonta, which raised the question of which form is prevailing across eukaryotes. The tight association of the hybrid TvTIM with ADP/ATP carriers (AAC) suggests that AAC may directly supply ATP for the protein import since ATP synthesis is limited in hydrogenosomes.
CONCLUSIONS: The hybrid TvTIM in hydrogenosomes represents an original structural solution that evolved for protein import when Δψ is negligible and remarkable example of evolutionary adaptation to an anaerobic lifestyle.
结果:我们报道了人类寄生虫阴道毛滴虫(TvTIM)的氢体中TIM的前所未有的修饰。我们表明,将含前序列的蛋白质导入氢脂质体基质是由杂合TIM22-TIM23复合物介导的,该复合物包括三个高度发散的核心成分,TvTim22、TvTim23和TvTim17样蛋白。TvTIM的混合特征由TvTim22和TvTim17/23的存在下划线,与小Tim伴侣(Tim9-10)相关联,在线粒体中已知有助于底物转移到TIM22复合物,以及与TIM23特异性ATP依赖性序列转位酶相关运动(PAM)的偶联。基于免疫共沉淀(coIP)和质谱的相互作用组重建表明,杂合TvTIM与旁系同源物的组成变化形成。132kDa纯化的TvTIM的单粒子电子显微镜显示存在单环的小Tims复合物,而线粒体TIM22复合物带有双胞胎小Tims六聚体。TvTIM目前是Opisthokonta之外的唯一可视化TIM,这提出了在真核生物中流行哪种形式的问题。杂合TvTIM与ADP/ATP载体(AAC)的紧密结合表明,AAC可以直接为蛋白质输入提供ATP,因为ATP的合成在氢原子体中受到限制。
结论:氢体中的杂种TvTIM代表了一种原始的结构解决方案,当ΔΦ可以忽略不计时,它就蛋白质输入而进化,并且是对厌氧生活方式的进化适应的显着例子。