Partial translocation of apocytochrome c across phospholipid bilayer

Abstract

Apocytochrome c (Apo c), heme-free cytochrome c, is synthesized in the cell cytoplasm without a cleaved signal sequence, and then translocated across the outer mitochondrial membrane. The interaction of Apo c with phospholipid vesicles has been studied as a model for understanding protein translocation across membrane. The translocation of Apo c across model membrane has been studied by using large unilamellar, trypsin-containing vesicles at neutral pH. It is observed with the electrophoretic experiment that the amount of Apo c decreases and that of a certain segment increase with time. The size of the segment is about 2.5 kDa, and larger than any segments that can be produced by complete tryptic digestion of Apo c. Also, the segment is labeled with hydrophobic probe partitioned in phospholipid bilayer. Thus, a part of Apo c is exposed to the internal trypsin of the vesicles and some segment is protected against tryptic digestion by phospholipid bilayer. The N-terminal amino acid of 2.5 kDa segment was proved to be methionine with dansylation method. So, the segment is the C-terminal part of Apo c. These results provide us the clue to understand the mechanism of Apo c-translocation.