title: Binding and release of cytochrome c in brain mitochondria is influenced by membrane potential and hydrophobic interactions with cardiolipin.
abstract: Factors influencing the release and anchorage of cytochrome c to the inner membrane of brain mitochondria have been investigated. Metabolic activity of mitochondria caused a decrease in the membrane potential delta psi(m), accompanied by detachment of the protein from the inner membrane. In a model system of cytochrome c reconstituted in cardiolipin (CL) liposomes, phosphate was used to breach the hydrophilic lipid-protein interactions. About 44% cytochrome c was removable when heart CL (80% 18:2n-6) was employed, whereas the remaining protein accounted for the tightly bound conformation characterized by hydrophobic lipid-protein interactions. Cytochrome c release from brain CL liposomes was higher compared to heart CL, consistent with lower polyunsaturated fatty acid content. The release was even higher with CL extracted from metabolically stressed mitochondria, exhibiting more saturated fatty acid profile compared to control (30% vs. 17%). Therefore, weakening of the hydrophobic interactions due to saturation of CL may account for the observed cytochrome c release from mitochondria following metabolic stress. Moreover, mitochondria enriched with polyunsaturated CL exhibited higher delta psi(m), compared to less unsaturated species, suggesting that CL fatty acid composition influences delta psi(m). Mitochondria incorporated exogenous cytochrome c without protease-sensitive factors or delta psi(m). The internalized protein anchored to the inner membrane without producing swelling, as monitored by forward and side light scattering, but produced delta psi(m) consumption, suggesting recovery of respiratory activity. The delta psi(m) decrease is ascribed to a selected mitochondrial population containing the incorporated cytochrome c.
authors.affiliation: Laboratory of Biochemistry, Department of Internal Medicine, University of Perugia, 06122, Perugia, Italy.
authors.forename: L
authors.initials: L
authors.lastname: Piccotti
authors.forename: M
authors.initials: M
authors.lastname: Buratta
authors.forename: S
authors.initials: S
authors.lastname: Giannini
authors.forename: P
authors.initials: P
authors.lastname: Gresele
authors.forename: R
authors.initials: R
authors.lastname: Roberti
authors.forename: L
authors.initials: L
authors.lastname: Corazzi
mesh_terms: D000818:Animals; D002308:Cardiolipins / Q000737:chemistry*; D002462:Cell Membrane / Q000502:physiology*; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000502:physiology*; D045304:Cytochromes c / Q000737:chemistry* / Q000378:metabolism*; D004789:Enzyme Activation; D057927:Hydrophobic and Hydrophilic Interactions; D008081:Liposomes / Q000737:chemistry; D008564:Membrane Potentials / Q000502:physiology*; D008928:Mitochondria / Q000502:physiology*; D018384:Oxidative Stress / Q000502:physiology; D010715:Phosphatidylglycerols / Q000737:chemistry; D011485:Protein Binding; D051381:Rats
chemical_list: D002308:Cardiolipins; D008081:Liposomes; D010715:Phosphatidylglycerols; D045304:Cytochromes c