Molecular Basis of Substrate Selectivity in the ADP/ATP Carrier

Abstract

The ADP/ATP carrier (AAC) is a membrane transporter that mediates the exchange of ADP and ATP across the mitochondrial inner membrane. During an exchange cycle, AAC switches between two conformational states, the cytoplasm-open state (c-state), and the matrix-open state (m-state). Our recent molecular dynamics simulations revealed spontaneous binding of ADP to the c-state AAC, and identified the unknown binding site for ADP as a pocket deeply positioned inside the lumen that forms through significant conformational changes of several basic residues in response to substrate binding. We also showed that ADP binding likely triggers AAC transition to the m-state by breaking a salt bridge network. The identified binding site has allowed us to explore substrate selectivity in AAC by simulating the “binding” of various ligands, e.g., AMP and Mg-ADP, to AAC. AMP does bind but is not transported by AAC, and Mg is known to have an inhibitory effect on AAC. However, the molecular details involved in these processes are largely unknown. Our results suggest that the presence of a minimum of two phosphate groups in their Mg-free form is absolutely necessary for proper binding and for initiating the structural changes required for activation of AAC. AMP and Mg-ADP cannot establish sufficient contact with the salt bridge ring at the bottom of AAC lumen, either due to lack of the beta-phosphate in AMP, or interference of Mg2+ with the phosphate groups in Mg-ADP. These results provide additional evidence for the ADP binding site characterized in our earlier study, and suggest a mechanism for substrate selectivity in AAC.