Specificity of the interaction of amino- and carboxy-terminal fragments of the mitochondrial precursor protein apocytochrome c with negatively charged phospholipids. A spin-label electron spin resonance study.

The contribution of the various regions of the mitochondrial precursor protein apocytochrome c to the interaction of the protein with phosphatidylserine dispersions has been studied with chemically and enzymatically prepared fragments of horse heart apocytochrome c and phospholipids spin-labeled at different positions of the sn-2 chain. Three amino-terminal heme-less peptides, two heme-containing amino-terminal fragments, one central fragment, and three carboxy-terminal fragments were studied. The electron spin resonance spectra of phospholipids spin-labeled at the C5 position of the fatty acid chain indicate that both amino-terminal and carboxy-terminal fragments of the apocytochrome c molecule cause a restriction of motion of the lipids, whereas the heme-containing peptides and protein have less effect. In addition, a second motionally more restricted lipid component, which is observed for apocytochrome c interacting with phosphatidylserine dispersions containing lipids spin-labeled at the C12 or C14 position [Görrissen, H., Marsh, D., Rietveld, A., & de Kruijff, B. (1986) Biochemistry 25, 2904-2910], was observed both on binding the carboxy-terminal fragments and on binding of the amino-terminal fragments of the precursor protein. Interestingly, even a small water-soluble peptide consisting of the 24 carboxy-terminal residues gave rise to a two-component spectrum, with an outer hyperfine splitting of the restricted lipid component of 59 G, indicating a considerable restriction of the chain motion. This suggests that both the carboxy- and amino-terminal parts of the protein penetrate into the center of the bilayer and cause a strong perturbation of the fatty acyl chain motion. The implications of these findings for the mechanism of apocytochrome c translocation across membranes are discussed.