Interaction of CO/NO with the Apoptosis-Inducing Cytochrome C -Cardiolipin Complex

Abstract

The interaction of mitochondrial cytochrome (cyt) c with cardiolipin (CL) is involved in the initial stages of apoptosis, leading to dissociation of the heme-Met80 bond and to a cyt c “second life” as a peroxidase.We show that in the presence of CL both carbon monoxide (CO) or nitric oxide (NO) bind to the cyt c heme with high affinity. Specifically, in cyt c-CL complexes the NO binding leads to a pentacoordinate heme In contrast, native cyt c involved in the respiratory chain does not bind CO. Nitrosyl adducts of ferrocyt c arepossible instead, though always hexacoordinate.Photodissociated CO from cyt c-CL-CO complexes gives rise to predominant bimolecular rebinding, while ∼20% is geminate in a ps timescale. This contrasts with Met80X mutant cyt c, where geminate recombination dominates due to the protein rigidity. These and other results are consistent with a CL anchorage model showing an acyl chain impaled in the protein: CL leads to changes in the protein conformation and flexibility, allowing ligands access to the heme. The CO affinity for cyt c-CL is high enough to envisage an antiapoptotic effect of nanomolar CO concentrations via inhibition of the cyt c peroxidase activity.Furthermore an unusually complex set of kinetic steps follows the initial NO binding to the cyt c-CL complex heme. NO binds via a scheme comparable to that described for cyt c′ and guanylate cyclase, the final product being NO attached to the proximal side of the heme. Features such as high yield of NO escape after dissociation, rapid dissociation of proximal histidine upon NO binding and its very fast binding after NO dissociation, the formation of hexacoordinate nitrosyl intermediates then indicate a remarkable mobility induced by CL on the proximal heme environment.