Affiliation: | (1) Timiryazev Institute of Plant Physiology RAS, Botanicheskaya 35, 127276 Moscow, Russia;(2) Department of Biology, Lomonosov Moscow State University, Vorobyovy Gory, 119899 Moscow, Russia |
Abstract: | Our previous investigations have established that Na+ translocation across the Tetraselmis viridis plasma membrane (PM) mediated by the primary ATP-driven Na+-pump, Na+-ATPase, is accompanied by H+ counter-transport [Y.V. Balnokin et al. (1999) FEBS Lett 462:402–406]. The hypothesis that the Na+-ATPase of T. viridis operates as an Na+/H+ exchanger is tested in the present work. The study of Na+ and H+ transport in PM vesicles isolated from T. viridis demonstrated that the membrane-permeant anion NO3– caused (i) an increase in ATP-driven Na+ uptake by the vesicles, (ii) an increase in (Na++ATP)-dependent vesicle lumen alkalization resulting from H+ efflux out of the vesicles and (iii) dissipation of electrical potential, , generated across the vesicle membrane by the Na+-ATPase. The (Na++ATP)-dependent lumen alkalization was not significantly affected by valinomycin, addition of which in the presence of K+ abolished at the vesicle membrane. The fact that the Na+-ATPase-mediated alkalization of the vesicle lumen is sustained in the absence of the transmembrane is consistent with a primary role of the Na+-ATPase in driving H+ outside the vesicles. The findings allowed us to conclude that the Na+-ATPase of T. viridis directly performs an exchange of Na+ for H+. Since the Na+-ATPase generates electric potential across the vesicle membrane, the transport stoichiometry is mNa+/nH+, where m>n.Abbreviations BTP Bis-Tris-Propane, 1,3-bis[tris(hydroxymethyl)methylamino]-propane - CCCP Carbonyl cyanide m-chlorophenylhydrazone - DTT Dithiothreitol - NCDC 2-Nitro-4-carboxyphenyl N,N-diphenylcarbamate - PMSF Phenylmethylsulfonyl fluoride - PM Plasma membrane |