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This article presents studies of light-dependent localization of enzyme activities in chloroplasts, which were performed with a combination of techniques that enable us to simultaneously follow both fast kinetics and slow transport processes. A modified low light: ratio imaging technique (LLRI) was used to study the light-dependent co-localization of dark-energy-dependent phosphorylation and dark-energy-independent ATP synthesis. This method enabled us to obtain good resolution of foci of enhanced phosphorylation or ATP synthesis. A technique to study light-dependent respiratory electron transport was also applied. In contrast to earlier studies, this technique was made much more sensitive by the use of antifreeze-treated Cyt b5 as a marker and by two-photon excitation. Images were obtained, and the light-dependent spatial localization of foci was studied with low photon fluxes. The results show that in chloroplasts in all the reactions investigated, dark-energy-dependent phosphorylation and ATP synthesis occur in foci of 5-15 microm diameter at the very periphery of photosynthetically competent chloroplasts.Parking Buying Guide
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