Relationships between rates of steady-state ATP synthesis and the magnitude of the proton-activity gradient across thylakoid membranes

Abstract

Rates of ATP synthesis by illuminated spinach chloroplast thylakoids in the steady state have been compared to the magnitude of the pH difference across the thylakoid membrane (ΔpH). The apparent dependence of the rate of photophosphorylation on ΔpH was not affected by the method used to vary these parameters. ΔpH generated by electron flow through each photosystem alone or through both photosystems was equivalent in its ability to drive ATP synthesis. The uncouplers, gramicidin and carbonylcyanide m-chlorophenylhydrazone, affected the relationship between phosphorylation and ΔpH in the same way as alteration in the rate of electron transport by changes in light intensity. Also, N,N′-dicyclohexylcarbodiimide, an inhibitor of phosphorylation, increased ΔpH as phosphorylation is inhibited. The Km of ADP for photophosphorylation was previously reported to increase when uncouplers were used to decrease the rate, but to decrease when the light intensity was lowered (Vinkler, C. (1981) Biochem. Biophys. Res. Commun. 99, 1095–1100). We show that higher uncoupler concentrations decrease the Km for ADP and provide an explanation for the increase in the Km caused by low uncoupler concentrations. These results are consistent with the concept that steady state ATP synthesis in thylakoids is driven by delocalized pH gradients.