Proton translocation of the bovine chromaffin-granule membrane

Abstract

Bovine chromaffin granules were lysed and their membranes resealed to give osmotically sensitive ‘ghosts’. These swell in the presence of salts and MgATP. It is shown that this is due to proton entry accompanied by anions. The rate of swelling depends on the anion present, but swelling is not limited to media containing permeant anions. It is quite marked in solutions of sulphates, phosphates and acetates. It is not uncoupler-sensitive, suggesting that at least one component of swelling is due to coupled proton and anion entry (non-electrogenic proton translocation). Direct measurements of transmembrane pH and potential gradients generated in the presence of MgATP shows that these are rapidly established in sucrose media, and are rather little affected by the presence of salts. They contribute roughly equally to the total protonmotive force. The potential gradient is establihsed very rapidly, but the pH gradient is generated over several minutes. The gradients are not completely dissipated by uncoupler, and it is shown that, in media containing sulphate but no permeant anion, sulphate can be taken up by the ‘ghosts’. There thus appear to be two mechanisms of proton translocation across the membrane, both dependent on ATP hydrolysis: an electrogenic transfer of protons, and proton movement linked to an anion transporter of broad specificity.