Structure, substrate binding and symmetry of the mitochondrial ADP/ATP carrier in its matrix-open state.

Abstract The mitochondrial ADP/ATP carrier (AAC) performs the first and last step in oxidative phosphorylation by exchanging, ADP and ATP across the mitochondrial inner membrane. Its optimal function has been shown to be dependent on cardiolipins (CLs), unique phospholipids located almost exclusively in the mitochondrial membrane. In addition, AAC exhibits an enthralling three-fold pseudo-symmetry, a unique feature of members of the SLC25 family. Recently, its conformation poised for binding of ATP was solved by X-ray crystallography, referred to as the matrix-state (m-state). Binding of the substrate leads to conformational changes that export of ATP to the mitochondrial intermembrane space. In this contribution, we investigate the influence of CLs on the structure, substrate-binding properties, and structural symmetry of the m-state, employing μs-scale molecular dynamics (MD) simulations. Our findings demonstrate that CLs play a minor stabilizing role on the AAC structure. The inter-domain salt-bridges and hydrogen bonds forming the cytoplasmic network and tyrosine braces, which ensure the integrity of the global AAC scaffold, highly benefit from the presence of CLs. Under these conditions, the carrier is found to be organized in a more compact structure in its interior, as revealed by analyses of the electrostatic potential, measure of the AAC cavity aperture, and the substrate-binding assays. Introducing a convenient structure-based symmetry metric, we quantified the structural three-fold pseudo-symmetry of AAC, not only for the crystallographic structure, but also for conformational states of the carrier explored in the MD simulations. Our results suggest that CLs moderately contribute to preserve the pseudo-symmetric structure of AAC. SIGNIFICANCE At both ends of oxidative phosphorylation, the mitochondrial ADP/ATP carrier (AAC) switches between two conformational states, the c- and m-states, to import and export nucleotides across the mitochondrial inner membrane. Its optimal function depends on cardiolipins, which stabilize the protein as it undergoes conformational transitions. Here, we assess how these lipids, ubiquitous to the mitochondrial membrane, modulate the structural stability, symmetry, and ATP-binding properties of the carrier in its m-state, and find that by strengthening inter-domain non-covalent interactions, they promote more compact conformations of the protein. In turn, the cardiolipin-induced structural rigidity of AAC regulates the number of conformations of ATP conducive for binding to the carrier. We also show that cardiolipins mildly preserve the three-fold pseudo-symmetry of the carrier.