Cytochrome c provokes the weakening of zwitterionic membranes as measured by force spectroscopy.

Cytochrome c (cyt c) is a small soluble protein from the intermembrane space of mitochondria. This protein is essential because it transfers electrons between two membrane complexes of the respiratory chain. In fact, during this transfer, the positively charged amino-acid residues surrounding the heme in the protein structure allow the cyt c to interact properly with the anionic part of other molecules: mainly the cardiolipin-rich membrane of mitochondria and respiratory complexes. We have previously shown that besides its interaction with anionic lipids, the cyt c is also able to cross neutral lipid membranes. In this work, with the help of AFM and punch-through experiments, we have measured the force required to penetrate the membrane in the fluid and in the gel phases with or without cyt c molecules. In the presence of cyt c molecules, the structures generated by the interaction with the protein were considerably weakened, which led to the desorption of the fluid bilayer and to a considerable loss of cohesion of the gel phase. These results show the usefulness of punch-through experiments in determining the changes of membrane properties in the presence of external agents.