β-Carotene Synthesis in Isolated Spinach Chloroplasts

Carefully isolated intact spinach chloroplasts virtually free of contamination of other organelles effectively form beta-carotene from NaH(14)CO(3) or [U-(14)C]-3-phosphoglycerate (PGA) under photosynthetic conditions. The photosynthate pool formed in chloroplasts from 1 to 2 millimolar [U-(14)C]-3-PGA or 3 to 6 millimolar NaH(14)CO(3) was fully sufficient to supply beta-carotene synthesis with intermediates for about 1 hour at maximal rates of about 20 nanomoles (14)C incorporated per milligram chlorophyll per hour. Fatty acid synthesis remains, under these circumstances, in linear dependence to substrate concentrations with far lower activity. Isotopic dilution of the beta-carotene synthesis by adding unlabeled glyceraldehyde 3-phosphate, dihydroxyacetone-P, 3-PGA, 2-PGA, phosphoenolpyruvate, pyruvate, respectively, may be interpreted as a direct substrate flow from photosynthetically fixed CO(2) to isopentenyl pyrophosphate synthesizing system. Unlabeled acetate did not dilute beta-carotene synthesis. Fatty acid synthesis acted similarly with unlabeled substrates; but it also was diluted by unlabeled acetate. These results indicate a tight linkage of photosynthetic carbon fixation and plastid isoprenoid synthesis.