IN-VITRO IMPORT OF THE RIESKE IRON-SULFUR PROTEIN BY TRYPANOSOME MITOCHONDRIA

Most of the proteins present in the mitochondrion are imported to that location from the cytosol. While this process has been studied extens ively in fungal and mammalian systems, little work has been done in ot her eukaryotic organisms. We are particularly interested in the Trypan osoma brucei system because this organism developmentally regulates mi tochondrial function during its life cycle and because one of the impo rted proteins lacks a conventional targeting sequence. We report here the development of an in vitro import system using crude trypanosome m itochondria and a nuclear encoded, mitochondrial protein. Import of th e Rieske iron-sulfur protein subunit of the cytochrome c reductase com plex requires a membrane potential, ATP, and a protein component on th e mitochondrial surface. The precursor protein is sequentially process ed to the mature form in two steps by peptidases that require divalent metal ions far activity. As in other eukaryotic systems, the first pr ocessing event occurs inside the inner membrane and is probably cataly zed by a matrix-processing protease. Surprisingly, the second processi ng activity is located outside the inner membrane. Both processing ste ps require ATP but are independent of a membrane potential. We suggest that the trypanosome iron-sulfur protein is imported along a ''conser vative sorting pathway'' but that the assembly mechanism of the reduct ase complex may be unique to trypanosomes.