Distinct domains control the addressing and the insertion of Bax into mitochondria

The translocation of Bax from the cytosol into the mitochondrial outer membrane is a central event during apoptosis. We report that beyond the addressing step, which involves its first alpha-helix (h alpha 1), the helices alpha 5 and alpha 6 (h alpha 5 alpha 6) are responsible for the insertion of Bax into mitochondrial outer membrane bilayer. The translocation of Bax to mitochondria is associated with specific changes in the conformation of the protein that are under the control of two prolines: Pro-13, which controls the unfolding of h alpha 1, and Pro-168, a proline located immediately before the hydrophobic carboxyl-terminal end (i.e. helix alpha 9, h alpha 9), which controls the disclosure of h alpha 5 alpha 6. An additional step, the disruption of an electrostatic bond formed between Asp-33 (h alpha 1) and Lys-64 (BH3), allows the mitochondria addressing of Bax. We conclude that, although the intramolecular interactions of h alpha 1 with the BH3 region control the addressing of Bax to mitochondria, the Pro-168 is involved in the control of its membrane insertion through h alpha 5 alpha 6.