The vacuolar H+-ATPase of lemon fruits is regulated by variable H+/ATP coupling and slip.
Lemon fruit tonoplasts, unlike those of seedling epicotyls, contain nitrate-insensitive H+-ATPase activity (Müller, M. L., Irkens-Kiesecker, U., Rubinstein, B., and Taiz, L. (1996) J. Biol. Chem. 271, 1916-1924). However, the degree of nitrate-insensitivity fluctuates during the course of the year with a seasonal frequency. Nitrate uncouples H+ pumping from ATP hydrolysis both in epicotyls and in nitrate-sensitive fruit V-ATPases. Neither bafilomycin nor oxidation cause uncoupling. The initial rate H+/ATP coupling ratios of epicotyl and the nitrate-sensitive fruit proton pumping activities are the same. However, the H+/ATP coupling ratio of the nitrate-insensitive fruit H+ pumping activity is lower than that of nitrate-sensitive and epicotyl V-ATPases. Several properties of the nitrate-insensitive H+-ATPase of the fruit indicate that it is a modified V-ATPase rather than a P-ATPase: 1) insensitivity to low concentrations of vanadate; 2) it is initially strongly uncoupled by nitrate, but regains coupling as catalysis proceeds; 3) both the nitrate-sensitive and nitrate-insensitive fruit H+-pumps have identical Km values for MgATP, and show similar pH-dependent slip and proton leakage rates. We conclude that the ability of the juice sac V-ATPase to build up steep pH gradients involves three factors: variable coupling, i.e. the ability to regain coupling under conditions that initially induce uncoupling; a low pH-dependent slip rate; the low proton permeability of the membrane.
Müller
M L
ML
Biology Department, Sinsheimer Laboratories, University of California, Santa Cruz, California 95064, USA.
Jensen
M
M
Taiz
L
L
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002384:Catalysis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005638:Fruit / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D018942:Macrolides; D009566:Nitrates / Q000494:pharmacology; D010084:Oxidation-Reduction; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011522:Protons; D025262:Vacuolar Proton-Translocating ATPases; D014638:Vanadates / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D018942:Macrolides; D009566:Nitrates; D011522:Protons; D014638:Vanadates; C040929:bafilomycin A1; D000255:Adenosine Triphosphate; D025262:Vacuolar Proton-Translocating ATPases; D006180:Proton-Translocating ATPases