Featured Article - April 2014
Short description: Structural and biophysical analyses of a membrane carrier ectodomain reveal a Ca2+-dependent switch that regulates ATP flux across the mitochondrial inner membrane.
Mitochondrial carriers mediate the transport of essential substrates across the inner membrane to the matrix space. The short calcium-binding mitochondrial carrier (SCaMC) is an abundant representative that regulates nucleotide levels in the matrix by transporting ATP in exchange for phosphate. SCaMC is unusual because it contains an N-terminal Ca2+-binding ectodomain (NTD) that potentially interacts with the transmembrane domain (TMD) to regulate activity. The Ca2+-dependent activation of SCaMC is not yet understood, mainly due to the absence of structural information on transport regulation by an ectodomain.
Chou and colleagues (PSI MPSbyNMR) solved the crystal structure of the human SCaMC NTD at 2.1-Å resolution by molecular replacement. The structure features two lobes, each containing two Ca2+-bound EF hands and in conformations similar to those seen in calmodulin, the ubiquitous calcium sensor. However, SCaMC NTD lacks the flexible linker present in calmodulin that allows the lobes in the latter to move freely and collapse into a compact conformation in complex with target sequences. Thus, the two lobes of the Ca2+-bound NTD are in a compact arrangement and interact extensively in the absence of a binding partner. An additional C-terminal helix in the NTD contains a conserved internal target sequence similar to those observed in several calmodulin complexes.
Since the NTD does not crystallize in the absence of calcium, the authors used NMR analyses to compare the apo and Ca2+-bound states. The experiments confirmed that the Ca2+-bound NTD adopts the same structure in solution as in the crystal and revealed that the NTD undergoes a major conformational transition upon binding calcium. Additionally, the apo form of the NTD is partially disordered and exhibits substantial local and global dynamics. Finally, the authors used NMR and surface plasmon resonance to determine whether the conformational switch influences interactions of the NTD with the TMD (reconstituted into proteoliposomes). Interestingly, the NTD and TMD only interact in the absence of calcium, with a dissociation constant of ∼250 μM.
Collectively, the data support a model in which the dynamic NTD interacts with the TMD in the absence of calcium to physically block ATP transport into the matrix. Calcium binding quenches the dynamics and relieves autoinhibition, suggesting a mechanism for SCaMC transporters.
Q. Yang et al. A self-sequestered calmodulin-like Ca2+ sensor of mitochondrial SCaMC carrier and its implication to Ca2+-dependent ATP-Mg/Pi transport.
Structure 22, 209-214 (2014). doi:10.1016/j.str.2013.10.018