Structure of the mitochondrial import gate reveals distinct preprotein paths.

The translocase of the outer mitochondrial membrane (TOM) is the main entry gate for proteins1-4 . Here we use cryo-electron microscopy to report the structure of the yeast TOM core complex5-9 at 3.8 Å resolution. The structure reveals the high-resolution architecture of the translocator consisting of two Tom40 β-barrel channels and α-helical transmembrane subunits, providing insight into critical features conserved in all eukaryotes1-3 . Each Tom40 β-barrel is surrounded by small Tom subunits, and tethered by two Tom22 and one phospholipid. The N-terminal extension of Tom40 forms a helix inside the channel; mutational analysis reveals its dual role in early and late steps in biogenesis of intermembrane-space proteins in cooperation with Tom5. Each Tom40 channel possesses two precursor exit sites: Tom22, Tom40 and Tom7 guide presequence-containing preproteins to the exit in the midst of the dimer, whereas Tom5 and the Tom40 N-extension guide presequence-less preproteins to the exit at the dimer periphery.