Ion Transport Across Chloroplast Inner Envelope Vesicles

Abstract

The ionic environment of chloroplasts is important for photosynthesis, nitrogen metabolism and sulfur metabolism, yet how this internal environment is regulated is still not known. Isolated chloroplast inner envelope membranes are competent for transport studies. These membranes can be manipulated to form vesicles of largely right side-out and inside-out orientation and both the intravesicular and extravesicular buffer contents can be controlled. The vesicles can be loaded with an ion-sensitive fluorophore to measure the initial rates of ion transport across the membranes using spectrofluorometric methods. We have made measurements on a proton-pumping ATPase which may be involved in forming a pH gradient to assist in the transport other ions across the chloroplast inner envelope. Several ions such a sulfate, glycolate, phosphate, iron and calcium have enhanced rates of transport when an inward-directed pH gradient is imposed. Calcium may actually move as a result of the potential gradient which is formed across the inner envelope. In addition both nitrite and bicarbonate movement is aided by the presence of a pH gradient, nitrite moving as nitrous acid and bicarbonate moving as carbon dioxide. Movement of the latter is greatly accelerated when carbonic anhydrase is present on both sides of the membrane. A summary of our results on the study of ion transport across the chloroplast inner envelope will be presented with respect to rates of transport, effectors, and inhibitors.