Reconstituted plant uncoupling mitochondrial protein allows for proton translocation via fatty acid cycling mechanism.
Potato and tomato plant uncoupling mitochondrial protein (PUMP) was reconstituted into liposomes, and K+ or H+ fluxes associated with fatty acid (FA)-induced ion movement were measured using fluorescent ion indicators potassium binding benzofuraneisophthalate and 6-methoxy-N-(3-sulfopropyl)-quinolinium. We suggest that PUMP, like its mammalian counterpart, the uncoupling protein of brown adipose tissue mitochondria (Garlid, K. D., Orosz, D. E., Modrianský, M., Vassanelli, S., and Jeek, P. (1996), J. Biol. Chem. 271, 2615-2702), allows for H+ translocation via a FA cycling mechanism. Reconstituted PUMP translocated anionic linoleic and heptylbenzoic acids, undecanesulfonate, and hexanesulfonate, but not phenylvaleric and abscisic acids or Cl-. Transport was inhibited by ATP and GDP. Internal acidification of protein-free liposomes by linoleic or heptylbenzoic acid indicated that H+ translocation occurs by FA flip-flopping across the lipid bilayer. However, addition of valinomycin after FA-initiated GDP-sensitive H+ efflux solely in proteoliposomes, indicating that influx of anionic FA via PUMP precedes a return of protonated FA carrying H+. Phenylvaleric acid, unable to flip-flop, was without effect. Kinetics of FA and undecanesulfonate uniport suggested the existence of an internal anion binding site. Exponential flux-voltage characteristics were also studied. We suggest that regulated uncoupling in plant mitochondria may be important during fruit ripening, senescence, and seed dormancy.
Jezek
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Department of Membrane Transport Biophysics, Institute of Physiology, Academy of Sciences, 14220 Prague 4, Czech Republic. jezek@sun1.biomed.cas.cz
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D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism*; D007473:Ion Channels; D007700:Kinetics; D018551:Solanum lycopersicum / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D024101:Mitochondrial Proteins; D011510:Proteolipids / Q000378:metabolism; D011522:Protons; D011198:Solanum tuberosum / Q000378:metabolism*; D013451:Sulfonic Acids / Q000378:metabolism; D000071256:Uncoupling Protein 1
D002352:Carrier Proteins; D005227:Fatty Acids; D007473:Ion Channels; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D011510:Proteolipids; D011522:Protons; D013451:Sulfonic Acids; D000071256:Uncoupling Protein 1; C020485:proteoliposomes