A covalent tandem dimer of the mitochondrial ADP/ATP carrier is functional in vivo.
The adenine nucleotide carrier, or Ancp, is an integral protein of the inner mitochondrial membrane. It is established that the inactive Ancp bound to one of its inhibitors (CATR or BA) is a dimer, but different contradictory models were proposed over the past years to describe the organization of the active Ancp. In order to decide in favor of a single model, it is necessary to establish the orientations of the N- and C-termini and thus the parity of the Ancp transmembrane segments (TMS). According to this, we have constructed a gene encoding a covalent tandem dimer of the Saccharomyces cerevisiae Anc2p and we demonstrate that it is stable and active in vivo as well as in vitro. The properties of the isolated dimer are strongly similar to those of the native Anc2p, as seen from nucleotide exchange and inhibitor binding experiments. We can therefore conclude that the native Anc2p has an even number of TMS and that the N- and C-terminal regions are exposed to the same cellular compartment. Furthermore, our results support the idea of a minimal dimeric functional organization of the Ancp in the mitochondrial membrane and we can suggest that TMS 1 of one monomer and TMS 6 of the other monomer in the native dimer are very close to each other.
Trézéguet
V
V
Laboratoire de Physiologie Moléculaire et Cellulaire, IBGC-CNRS, 1 rue Camille Saint-Saëns, 33077, Bordeaux cedex, France. vero.tezeguet@ibgc.u-bordeaux2.fr
Le Saux
A
A
David
C
C
Gourdet
C
C
Fiore
C
C
Dianoux
A
A
Brandolin
G
G
Lauquin
G J
GJ
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D019281:Dimerization; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D000226:Mitochondrial ADP, ATP Translocases / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins
D005656:Fungal Proteins; D009687:Nuclear Proteins; C112225:PET9 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D000226:Mitochondrial ADP, ATP Translocases