Alkalization of the chloroplast stroma caused by light-dependent proton flux into the thylakoid space

Abstract

1. From the uptake of dimethyloxazolidinedione and of methylamine into the sucrose-impermeable space of intact spinach chloroplasts as measured by siliconlayer filtering centrifugation and from estimation of the size of the thylakoid space by planimetry of electron micrographs the pH in the stroma and in the thylakoid space is evaluated. The reliability of the method is checked. 2. Illumination causes an alkalization in the stroma and an acidification in the thylakoid space, with a ΔpH of about 2.5 between the two spaces, reflecting light-dependent proton transport across the thylakoid membrane. By addition of m-chlorocarbonyl cyanide phenylhydrazone or of nigericin these pH changes are reversed to the corresponding dark values. 3. Beside this there is also some light-dependent proton movement from the stroma across the inner membrane into the external space. 4. The pH in the stroma and in the thylakoid space of illuminated chloroplasts depends on the pH in the medium, whereas the ΔpH across the thylakoid membrane is almost independent from this. 5. From comparison of the pH changes in the stroma and in the thylakoid space it appears that in the thylakoid space there is a buffer with p K 5.5 and in the stroma with p K 6.8 and that the buffer concentration in the thylakoid space is about four times higher than in the stroma. 6. It is discussed that the light-dependent alkalization in the chloroplast stroma may be involved in the regulation of CO 2 fixation at the fructose diphosphatase step.