Effects of Magnesium on Intact Chloroplasts : II. CATION SPECIFICITY AND INVOLVEMENT OF THE ENVELOPE ATPase IN (SODIUM) POTASSIUM/PROTON EXCHANGE ACROSS THE ENVELOPE

Abstract

Addition of exogenous Mg(2+) (2 millimolar) to illuminated intact spinach (Spinacia oleracea L.) chloroplasts caused acidification of the stroma and a 20% decrease in stromal K(+). Addition of K(+) (10-50 millimolar) reversed both stromal acidification and K(+) efflux from the chloroplast caused by Mg(2+). These data suggested that Mg(2+) induced reversible H(+)/K(+) fluxes across the chloroplast envelope. Ca(2+) and Mn(2+) (2 millimolar) were as effective as 4 millimolar Mg(2+) in causing K(+) efflux from chloroplasts and inhibition of O(2) evolution. In contrast, 10 millimolar Ba(2+) induced only a small amount of inhibition. The lack of strong inhibition by Ba(2+) indicated that the effects of divalent cations such as Mg(2+) cannot be attributed to generalized electrostatic interactions of the cation with the chloroplast envelope. With the chloroplasts used in this study, stromal acidification caused by 2 millimolar Mg(2+) was small (0.07 to 0.15 pH units), but sufficient to account for the inhibition of O(2) evolution (43%) induced by Mg(2+).A variety of ATPase inhibitors were tested for effects on Mg(2+)-induced H(+)/K(+) fluxes. Oligomycin was the only ATPase inhibitor which specifically inhibited photosynthesis in the presence of Mg(2+) + K(+), but had little or no effect in the absence of these cations. In the presence of oligomycin, much higher concentrations (50 millimolar) of exogenous K(+) were required to reverse Mg(2+)-induced acidification and inhibition of O(2) evolution than in its absence. Oligomycin (in the absence of divalent cations) increased the inhibition of photosynthesis caused by sodium acetate, which acts by causing stromal acidification. In addition, the chloroplast envelope ATPase was inhibited partially (45%) by oligomycin. These results suggested that H(+) fluxes across the chloroplast envelope are regulated by two mechanisms: (a) an active, oligomycin-sensitive H(+) efflux and (b) a reversible, Mg(2+)-dependent, oligomycin-insensitive H(+)/K(+) exchange.