Effect of high KCl concentrations on membrane-localized metastable proton buffering domains in thylakoids

Recent work showed that chloroplast thylakoid membranes stored in 100 mM KCl-containing media have delocalized energy coupling consistent with a rapid equilibration of the proton gradient between the proton-producing redox steps and the lumen bulk phase (Beard and Dilley 1986). Thylakoids stored in low salt media showed localized energy coupling. A related thylakoid membrane property is the occurrence of sequestered, metastable, acidic domains, associated with pK _a 7.5 amine groups. For low salt-stored membranes the domain protons appear to be in the direct (localized) diffusion pathway of protons involved in energizing ATP formation, whereas in thylakoids stored in high KCl, domain protons equilibrated with the lumen during the development of the ATP energization threshold (Theg et al. 1988). This work tested whether the 100 mM KCl storage treatment did or did not cause the dissipation of the metastable acidic domain protons in the dark, storage period. By three criteria, it was found that the 100 mM KCl storage treatment had only a slight tendency to dissipate the acidic domain protons into alkaline media under dark conditions. Storage in KCl does not cause the dissipation of the acidic domains in the dark, but allows domain protons to equilibrate with the lumen after the redox system begins turning over, but before the ATP energization threshold pH is reached. These results must be considered in models of how the thylakoid structure can accommodate metastable acidic domains and how such domain protons diffuse to the CF₀-CF₁ complexes in energy coupling.Abbreviations PSII photosystem 2 - HEPES N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid - MES 2(N-morpholine) ethanesulfonic acid - Taps N-tris (hydroxymethyl)-methyl-3-amino-propanesulfonic acid - EPPS N-(2-hydroxyethyl) piperazine-N-3-propanesulfonic acid - gram D gramicidin D This research supported in part by grants from the USDA and NSF.