Sulfite both stimulates and inhibits the yeast vacuolar H(+)-ATPase.
The yeast vacuolar H(+)-ATPase (V-ATPase) exhibits nonlinear hydrolysis kinetics, i.e. an initial rapid rate followed by a slower, steadily declining rate. Sulfite (50-100 mM) stimulates the yeast V-ATPase specifically during the latter period. Sulfite activation has been observed for the F-ATPases and archaebacterial ATPases and is thought to be caused by the release of tightly bound nucleotide at the catalytic site. However, turnover-dependent inactivation of the yeast V-ATPase, and sulfite stimulation, were only observed at MgATP concentrations > 1.0 mM. Below 1.0 mM MgATP, the hydrolysis time course was linear, and sulfite was inhibitory. The inhibition during the initial phase and the stimulation during the later phase of the time course could be accounted for by a 5.5-fold sulfite-induced increase in the apparent Km, and a small increase in the apparent Vmax. Sulfite also protected the enzyme against inhibition by cold inactivation and by dicyclohexylcarbodiimide but not by bafilomycin. Sulfite stimulation during the later phase was antagonized by delta mu H+, particularly by delta pH. In contrast to its effects on hydrolysis, sulfite inhibited the formation of a pH gradient at all times and failed to enhance the membrane potential even when delta pH was collapsed by nigericin. These results indicate that sulfite partially uncouples hydrolysis from proton transport in a way that preserves regulation by delta mu H+.
Kibak
H
H
Department of Biology, Sinsheimer Laboratories, University of California, Santa Cruz 95064.
Van Eeckhout
D
D
Cutler
T
T
Taiz
S L
SL
Taiz
L
L
D000255:Adenosine Triphosphate / Q000378:metabolism; D001693:Biological Transport, Active / Q000187:drug effects; D003080:Cold Temperature; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D013447:Sulfites / Q000494:pharmacology*; D014617:Vacuoles / Q000201:enzymology*
D013447:Sulfites; D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases; D008274:Magnesium