Article
Defining the Apoptotic Trigger: the Interaction of Cytochrome c and Cardiolipin
The Journal of Biological Chemistry
(2015)
Abstract
The interaction between cytochrome c and the anionic lipid cardiolipin has been proposed as a primary event in the apoptotic signaling cascade. Numerous studies that have examined the interaction of cytochrome c with cardiolipin embedded in a variety of model phospholipid membranes have suggested that partial unfolding of the protein is a precursor to the apoptotic response. However, these studies lacked site resolution and used model systems with negligible or a positive membrane curvature, which is distinct from the large negative curvature of the invaginations of the inner mitochondrial membrane where cytochrome c resides. We have used reverse micelle encapsulation to mimic the potential effects of confinement on the interaction of cytochrome c with cardiolipin. Encapsulation of oxidized horse cytochrome c in 1-decanoyl-rac-glycerol/lauryldimethylamine-N-oxide/hexanol reverse micelles prepared in pentane yields NMR spectra essentially identical to the protein in free aqueous solution. The structure of encapsulated ferricytochrome c was determined to high precision (<r.m.s. deviation>bb ∼ 0.23 Å) using NMR-based methods and is closely similar to the cryogenic crystal structure (<r.m.s. deviation>bb ∼ 1.2 Å). Incorporation of cardiolipin into the reverse micelle surfactant shell causes localized chemical shift perturbations of the encapsulated protein, providing the first view of the cardiolipin/cytochrome c interaction interface at atomic resolution. Three distinct sites of interaction are detected: the so-called A- and L-sites, plus a previously undocumented interaction centered on residues Phe-36, Gly-37, Thr-58, Trp-59, and Lys-60. Importantly, in distinct contrast to earlier studies of this interaction, the protein is not significantly disturbed by the binding of cardiolipin in the context of the reverse micelle.
Keywords
- apoptosis,
- cardiolipin,
- cytochrome c,
- lipid-protein interaction,
- nuclear magnetic resonance (NMR),
- protein folding,
- protein stability,
- confined space,
- protein hydration
Disciplines
Publication Date
December 25, 2015
DOI
10.1074/jbc.M115.689406
Citation Information
Evan S. O'Brien, Nathaniel V. Nucci, Brian Fuglestad, Cecilia Tommos, et al.. "Defining the Apoptotic Trigger: the Interaction of Cytochrome c and Cardiolipin" The Journal of Biological Chemistry Vol. 290 Iss. 52 (2015) p. 30879 - 30887 ISSN: 0021-9258 Available at: http://works.bepress.com/nathaniel-nucci/1/