Apocytochrome c induces pH-dependent vesicle fusion

Area of Science:

• Biochemistry
• Biophysics
• Membrane Biology

Background:

• Cytochrome c is a key component of the respiratory chain.
• Apocytochrome c is the heme-free precursor to cytochrome c.
• Protein-lipid interactions are crucial for membrane fusion and protein translocation.

Purpose of the Study:

• To investigate the ability of apocytochrome c and cytochrome c to induce fusion of small unilamellar vesicles.
• To examine the influence of negatively charged lipids and pH on vesicle fusion.
• To understand the conformational changes of apocytochrome c during vesicle interaction.

Main Methods:

• Fusion assay using energy transfer between fluorescent lipid probes.
• Vesicles composed of phosphatidylcholine (PC) with varying amounts of negatively charged lipids (phosphatidylserine, cardiolipin, phosphatidylglycerol, phosphatidylinositol).
• Spectroscopic analysis of apocytochrome c secondary structure changes.

Main Results:

• Both apocytochrome c and cytochrome c mediated vesicle fusion in a concentration- and pH-dependent manner.
• Apocytochrome c induced fusion over a broader pH range and was more potent than cytochrome c.
• Maximal fusion occurred at acidic pH (3-4), inducing conformational changes in apocytochrome c from random coil to alpha-helix in the presence of vesicles.
• Fusion increased with higher phosphatidylserine content and was also observed in pure PC vesicles.

Conclusions:

• The pH dependence of fusion is attributed to changes in net charge on both vesicles and proteins.
• Apocytochrome c's ability to induce fusion in various vesicle compositions may relate to its biological translocation across mitochondrial membranes.
• Protein-induced membrane fusion is influenced by lipid composition, protein charge, and conformational changes.