Molecular aspects of the adenine nucleotide carrier from mitochondria

Abstract

The ADP/ATP carrier (AAC) of mitochondria is a functionally central and characteristic component of the eukaryotic cell. By linking the thermodynamically divergent metabolites in the intra- and extramitochondrial compartments, it had to evolve with the emergence of the eukaryotic cell. Because of a number of unique properties, the AAC provided advanced insight into the molecular basis of solute transport through biomembrane carriers. With highly specific and unusually large substrates, ADP and ATP, and with high-affinity inhibitors binding selectively either from the inside or the outside, the first molecular demonstration of the single-binding-center gated pore mechanism was made. This framework can only partially be interpreted with the available yet rapidly increasing structural information on the AAC. The primary structure, first established for the AAC from beef heart mitochondria, showed a relatively wide distribution of hydrophilic residues which permits assignment of only two hydrophobic transmembrane stretches. However, a striking tripartition of the primary structure into about three 100-residue-long domains allows a more significant assignment of transmembrane elements. With alignment of these three domains for maximum conservation of structurally critical residues, each domain can be assigned to have two transmembrane α elements between 18 and 22 residues long. The interdomain homology between these α regions is low. The central regions flanked by these helices contain most of the polar residues and are significantly interdomain conserved. With lysine probes the central regions are assigned to the matrix side (m-side) and the two connecting regions as well as C and N termini to the cytosolic side (c-side). Out of the central regions a loop is assumed to protrude through the membrane, probably for lining the translocation channel. This localization of a major protein mass within the membrane agrees with hydrodynamic evidence, the carrier being an oblate ellipsoid with only about 50 Å along the short axis. In accordance, the loops of domains 2 and 3 are affinity labeled by azido-ADP or azido-atractylate. Primary structures of AAC from other sources (fungi, plants) also exhibit the tripartition. The interdomain conserved residues are also interspecies conserved, thus showing that they are essential. These repeat domains have probably evolved from a common gene coding for about 100 residues. Isoforms of the AAC exist, as shown by primary structure analysis of human cDNA libraries from different organs. Three different isoforms are identified in human organs. These isoforms may have developed in accordance with specific metabolic requirements of various tissues for ADP/ ATP exchange.