title: In vitro analyses of mitochondrial ATP/phosphate carriers from Arabidopsis thaliana revealed unexpected Ca(2+)-effects.
abstract: BACKGROUND
  Adenine nucleotide/phosphate carriers (APCs) from mammals and yeast are commonly known to adapt the mitochondrial adenine nucleotide pool in accordance to cellular demands. They catalyze adenine nucleotide--particularly ATP-Mg--and phosphate exchange and their activity is regulated by calcium. Our current knowledge about corresponding proteins from plants is comparably limited. Recently, the three putative APCs from Arabidopsis thaliana were shown to restore the specific growth phenotype of APC yeast loss-of-function mutants and to interact with calcium via their N-terminal EF--hand motifs in vitro. In this study, we performed biochemical characterization of all three APC isoforms from A. thaliana to gain further insights into their functional properties.
  
  
  RESULTS
  Recombinant plant APCs were functionally reconstituted into liposomes and their biochemical characteristics were determined by transport measurements using radiolabeled substrates. All three plant APCs were capable of ATP, ADP and phosphate exchange, however, high preference for ATP-Mg, as shown for orthologous carriers, was not detectable. By contrast, the obtained data suggest that in the liposomal system the plant APCs rather favor ATP-Ca as substrate. Moreover, investigation of a representative mutant APC protein revealed that the observed calcium effects on ATP transport did not primarily/essentially involve Ca(2+)-binding to the EF-hand motifs in the N-terminal domain of the carrier.
  
  
  CONCLUSIONS
  Biochemical characteristics suggest that plant APCs can mediate net transport of adenine nucleotides and hence, like their pendants from animals and yeast, might be involved in the alteration of the mitochondrial adenine nucleotide pool. Although, ATP-Ca was identified as an apparent import substrate of plant APCs in vitro it is arguable whether ATP-Ca formation and thus the corresponding transport can take place in vivo.
authors.affiliation: Cellular Physiology/Membrane Transport, University of Kaiserslautern, 67653, Kaiserslautern, Germany. anlorenz@rhrk.uni-kl.de.
authors.forename: André
authors.initials: A
authors.lastname: Lorenz
authors.affiliation: Cellular Physiology/Membrane Transport, University of Kaiserslautern, 67653, Kaiserslautern, Germany. lorenz-melanie@gmx.net.
authors.forename: Melanie
authors.initials: M
authors.lastname: Lorenz
authors.affiliation: Department of Biology I, Botany, LMU Munich, Großhaderner Str. 2, D-82152, Planegg-Martinsried, Germany. vothknecht@lrz.uni-muenchen.de.
authors.forename: Ute C
authors.initials: UC
authors.lastname: Vothknecht
authors.affiliation: Plant Physiology, University of Kaiserslautern, 67653, Kaiserslautern, Germany. witzsand@rhrk.uni-kl.de.
authors.forename: Sandra
authors.initials: S
authors.lastname: Niopek-Witz
authors.affiliation: Plant Physiology, University of Kaiserslautern, 67653, Kaiserslautern, Germany. neuhaus@rhrk.uni-kl.de.
authors.forename: H Ekkehard
authors.initials: HE
authors.lastname: Neuhaus
authors.affiliation: Cellular Physiology/Membrane Transport, University of Kaiserslautern, 67653, Kaiserslautern, Germany. haferk@rhrk.uni-kl.de.
authors.forename: Ilka
authors.initials: I
authors.lastname: Haferkamp
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D017920:Antiporters / Q000378:metabolism; D017360:Arabidopsis / Q000187:drug effects / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D002118:Calcium / Q000494:pharmacology*; D002413:Cations, Divalent / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D006801:Humans; D008274:Magnesium / Q000494:pharmacology; D024101:Mitochondrial Proteins / Q000378:metabolism*; D028061:Phosphate Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011995:Recombination, Genetic / Q000235:genetics; D013997:Time Factors
chemical_list: D017920:Antiporters; D029681:Arabidopsis Proteins; D002413:Cations, Divalent; D024101:Mitochondrial Proteins; D028061:Phosphate Transport Proteins; D004533:Egtazic Acid; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D008274:Magnesium; D002118:Calcium