METABOLITE LEVELS IN THE CHLOROPLAST AND EXTRACHLOROPLAST COMPARTMENTS OF BARLEY LEAF PROTOPLASTS DURING THE INITIAL PHASE OF PHOTOSYNTHETIC INDUCTION

Metabolite levels were determined in the chloroplast and extrachloroplast compartments of barley protoplasts during photosynthetic induction using rapid fractionation by membrane filtration. This method allowed studies with a high time resolution the first determination of subcellular metabolite content bring made after only 0.3 s. Upon illumination, dark-adapted protoplasts exhibited a 1 min lag phase prior to commencement of oxygen evolution, and the maximum rate was reached after 4 to 5 min. In contrast to oxygen evolution, the ATP/ADP ratio in the chloroplasts increased from 1 to 2 within 0.5 s and reached a maximum of about 5 after 2 s. There was a dramatic increase in the extrachloroplastic ATP/ADP ratio within a few seconds, reaching a maximum after about 15 s. During the initial phase of photosynthetic induction, the subcellular ATP/ADP ratios were very similar in photorespiratory (low CO,) and non-photorespiratory (high CO,) conditions. The ATP/ADP ratios in both the chloroplast and extrachloroplast compartments remained high until photosynthetic oxygen evolution started and then decreased when the photosynthetic rate reached its maximum. In steady-state photosynthesis the subcellular ATP/ADP ratios were considerably higher under photorespiratory conditions as compared to non-photorespiratory conditions. During the initial phase of photosynthetic induction, 3-phosphoglycerate decreased and triose phosphates increased both in the chloroplast and extrachloroplast compartments. The changes in these metabolites are consistent with a 3-phosphoglycerate/triose phosphate shuttle using the phosphate translocator as the means to supply ATP to the cytosol during photosynthetic induction.