The membrane sector of vacuolar H(+)-ATPase by itself is impermeable to protons

The sensitivity for vanadate of proton uptake activities of isolated chromaffin granules and yeast vacuoles was investigated. About 0.5 mM vanadate caused 50% inhibition of ATP-dependent proton uptake activity in both membranes. The proton conductivity across chromaffin granule membranes and yeast vacuoles was assayed in the presence and absence of the catalytic sectors of their respective V-ATPases. Removal of the catalytic sectors by cold treatment in the presence of MgATP did not change the proton conductivity of the membranes. Similar results were obtained with yeast vacuoles of mutants that did not assemble the catalytic sector of the enzyme. These results are in contrast to the effect of removing the catalytic sectors of F-ATPases from various sources. The mechanistic and biological significance of the difference in the behavior of the two families of proton pumps is discussed.