Characterization of the Tim17 associated protein, Tim50, in Trypanosoma brucei

Abstract

Trypanosoma brucei causes a fatal disease in humans and domestic animals known as African trypanosomaisis. T. brucei possesses a single, tubular mitochondrion with many unique characteristics. Similar to other eukaryotes, most of the mitochondrial proteins in T. brucei are encoded in the nucleus and must be transported into the mitochondrion. However, mitochondrial protein import machinery in T. brucei is poorly understood. In eukaryotes, proteins are imported into mitochondria post‐transcriptionally via receptor/translocase complexes located on mitochondrial outer and inner membranes known as TOM and TIMs. In T. brucei, only Tim17 has been characterized from the TIM complex. Recently we have identified Tim50 in this parasitic protozoan. T. brucei Tim50 (TbTim50) protein sequence showed 16–18% identity and 28–30% similarity with Tim50 from other eukaryotes. The putative Tim50 possesses an N‐terminal mitochondrial targeting sequence. Expression of the C‐terminal HA‐tagged TbTim50 in T. brucei followed by sub‐cellular fractionation and western blot analysis revealed that the expressed protein is enriched in the mitochondrial fraction. Anti‐HA antibody was able to co‐immunoprecipitate Tim17 from T. brucei mitochondrial lysate, leading us to conclude TbTim50 is associated with Tim17. This work was supported by NIH grants SC1GM081146 and 2T32HL007737‐16.