title: Functional complementation analyses reveal that the single PRAT family protein of trypanosoma brucei is a divergent homolog of Tim17 in saccharomyces cerevisiae.
abstract: Trypanosoma brucei, a parasitic protozoan that causes African trypanosomiasis, possesses a single member of the presequence and amino acid transporter (PRAT) protein family, which is referred to as TbTim17. In contrast, three homologous proteins, ScTim23, ScTim17, and ScTim22, are found in Saccharomyces cerevisiae and higher eukaryotes. Here, we show that TbTim17 cannot rescue Tim17, Tim23, or Tim22 mutants of S. cerevisiae. We expressed S. cerevisiae Tim23, Tim17, and Tim22 in T. brucei. These heterologous proteins were properly imported into mitochondria in the parasite. Further analysis revealed that although ScTim23 and ScTim17 were integrated into the mitochondrial inner membrane and assembled into a protein complex similar in size to TbTim17, only ScTim17 was stably associated with TbTim17. In contrast, ScTim22 existed as a protease-sensitive soluble protein in the T. brucei mitochondrion. In addition, the growth defect caused by TbTim17 knockdown in T. brucei was partially restored by the expression of ScTim17 but not by the expression of either ScTim23 or ScTim22, whereas the expression of TbTim17 fully complemented the growth defect caused by TbTim17 knockdown, as anticipated. Similar to the findings for cell growth, the defect in the import of mitochondrial proteins due to depletion of TbTim17 was in part restored by the expression of ScTim17 but was not complemented by the expression of either ScTim23 or ScTim22. Together, these results suggest that TbTim17 is divergent compared to ScTim23 but that its function is closer to that of ScTim17. In addition, ScTim22 could not be sorted properly in the T. brucei mitochondrion and thus failed to complement the function of TbTim17.
authors.affiliation: Department of Microbiology and Immunology, Meharry Medical College, Nashville, Tennessee, USA.
authors.forename: Ebony
authors.initials: E
authors.lastname: Weems
authors.affiliation: Department of Microbiology and Immunology, Meharry Medical College, Nashville, Tennessee, USA.
authors.forename: Ujjal K
authors.initials: UK
authors.lastname: Singha
authors.affiliation: Department of Microbiology and Immunology, Meharry Medical College, Nashville, Tennessee, USA.
authors.forename: VaNae
authors.initials: V
authors.lastname: Hamilton
authors.affiliation: Department of Microbiology and Immunology, Meharry Medical College, Nashville, Tennessee, USA.
authors.forename: Joseph T
authors.initials: JT
authors.lastname: Smith
authors.affiliation: Adolf Butenandt Institute, Physiological Chemistry, University of Munich, Munich, Germany.
authors.forename: Karin
authors.initials: K
authors.lastname: Waegemann
authors.affiliation: Adolf Butenandt Institute, Physiological Chemistry, University of Munich, Munich, Germany.
authors.forename: Dejana
authors.initials: D
authors.lastname: Mokranjac
authors.affiliation: Department of Microbiology and Immunology, Meharry Medical College, Nashville, Tennessee, USA mchaudhuri@mmc.edu.
authors.forename: Minu
authors.initials: M
authors.lastname: Chaudhuri
mesh_terms: D000595:Amino Acid Sequence; D005816:Genetic Complementation Test; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D033681:Mitochondrial Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D008969:Molecular Sequence Data; D011485:Protein Binding; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014346:Trypanosoma brucei brucei / Q000235:genetics / Q000378:metabolism*
chemical_list: D026901:Membrane Transport Proteins; D033681:Mitochondrial Membrane Transport Proteins; D000090622:Mitochondrial Precursor Protein Import Complex Proteins; D015800:Protozoan Proteins; D029701:Saccharomyces cerevisiae Proteins; C091191:TIM17 protein, S cerevisiae; C102549:TIM22 protein, S cerevisiae; C083081:TIM23 protein, S cerevisiae