title: The yeast mitochondrial ADP/ATP carrier functions as a monomer in mitochondrial membranes.
abstract: Mitochondrial carriers are believed widely to be dimers both in structure and function. However, the structural fold is a barrel of six transmembrane alpha-helices without an obvious dimerisation interface. Here, we show by negative dominance studies that the yeast mitochondrial ADP/ATP carrier 2 from Saccharomyces cerevisiae (AAC2) is functional as a monomer in the mitochondrial membrane. Adenine nucleotide transport by wild-type AAC2 is inhibited by the sulfhydryl reagent 2-sulfonatoethyl-methanethiosulfonate (MTSES), whereas the activity of a mutant AAC2, devoid of cysteines, is unaffected. Wild-type and cysteine-less AAC2 were coexpressed in different molar ratios in yeast mitochondrial membranes. After addition of MTSES the residual transport activity correlated linearly with the fraction of cysteine-less carrier present in the membranes, and so the two versions functioned independently of each other. Also, the cysteine-less and wild-type carriers were purified separately, mixed in defined ratios and reconstituted into liposomes. Again, the residual transport activity in the presence of MTSES depended linearly on the amount of cysteine-less carrier. Thus, the entire transport cycle for ADP/ATP exchange is carried out by the monomer.
authors.affiliation: Medical Research Council, Dunn Human Nutrition Unit, Hills Road, Cambridge CB2 2XY, United Kingdom.
authors.forename: Lisa
authors.initials: L
authors.lastname: Bamber
authors.forename: Marilyn
authors.initials: M
authors.lastname: Harding
authors.forename: Magnus
authors.initials: M
authors.lastname: Monné
authors.forename: Dirk-Jan
authors.initials: DJ
authors.lastname: Slotboom
authors.forename: Edmund R S
authors.initials: ER
authors.lastname: Kunji
mesh_terms: D000227:Adenine Nucleotides / Q000378:metabolism; D007700:Kinetics; D008698:Mesylates / Q000494:pharmacology; D000226:Mitochondrial ADP, ATP Translocases / Q000737:chemistry* / Q000378:metabolism*; D051336:Mitochondrial Membranes; D011487:Protein Conformation; D020836:Protein Structure, Quaternary; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism*; D013439:Sulfhydryl Reagents / Q000494:pharmacology
chemical_list: D000227:Adenine Nucleotides; D008698:Mesylates; C112225:PET9 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D013439:Sulfhydryl Reagents; C109153:(2-sulfonatoethyl)methanethiosulfonate; D000226:Mitochondrial ADP, ATP Translocases