Regulation of the reconstituted chloroplast phosphate translocator by an H+ gradient

This report describes the influence of ΔpH on the transport of phosphate, triose phosphate and 3-phosphoglycerate catalyzed by the phosphate translocator in a reconstituted system. The H⁺ gradient across the liposome membrane is adjusted by the addition of external buffer solution and maintained for several minutes. The following results are obtained: (1) An inward directed H⁺ gradient leads to an increase of 3-phosphoglycerate transport and to a decrease of phosphate and triose phosphate transport. (2) An H⁺ gradient in the opposite direction results in a restriction of 3-phosphoglycerate influx whereas the influx of phosphate and triose phosphate is enhanced. (3) The magnitude of the pH effect depends on the internal substrate. Compared to the homoexchange mode, the effect of applied ΔpH is more pronounced in the heteroexchange mode. (4) Transport of phosphate and 3-phosphoglycerate is influenced by ΔpH in a different manner. In the case of phosphate and triose phosphate transport the observed effects are associated with changes in the apparent K_m values whereas in the case of 3-phosphoglycerate transport the application of a pH gradient is linked to a change of V_max. (5) In competition experiments with both substrates in the external medium, ΔpH influences the effect of phosphate as a competitive inhibitor of 3-phosphoglycerate transport whereas the effect of 3-phosphoglycerate on phosphate transport is not affected by a pH gradient. (6) The measured apparent K_m and V_max values under the influence of ΔpH can be used for the calculation of substrate fluxes across the envelope during illumination. It can be demonstrated that the increase of stromal pH in the light gives rise to a considerable change in the ratio of the substrates transported. Under conditions without pH gradient, the species transported out is mainly 3-phosphoglycerate and the species transported in is mainly triose phosphate. These fluxes are reversed when a pH gradient is applied (light conditions).