SWEET-PEPPER PLASTIDS - ENZYMATIC EQUIPMENT, CHARACTERIZATION OF THE PLASTIDIC OXIDATIVE PENTOSE-PHOSPHATE PATHWAY, AND TRANSPORT OF PHOSPHORYLATED INTERMEDIATES ACROSS THE ENVELOPE MEMBRANE

Chloroplasts or chromoplasts were purified from sweet-pepper (Capsicum annuum L. cv. Wonder) fruits and analysed with respect to enzymic equ ipment, the transport properties across the envelope membrane, and for the presence of a functional oxidative pentose-phosphate pathway (OPP P). It was demonstrated that both types of plastid contain enzyme acti vities that allow glycolysis and OPPP. During the developmental conver sion from chloroplasts to chromoplasts the activities of enzymes catal ysing potentially rate-limiting reactions in glycolysis increased cons iderably. Most enzyme activities involved in the plastidic OPPP stayed constant or decreased during ripening, but transaldolase activity inc reased by more than 500%. To analyse whether pepper fruit chromoplasts are able to use exogenously supplied carbohydrates for the OPPP we me asured the rate of (CO2)-C-14 release after application of radioactive ly labelled precursors. Isolated pepper fruit chromoplasts used exogen ously supplied [(UC)-C-14]glucose-6-phosphate (Glc6P) as a precursor f or the OPPP. The metabolic flux through this pathway was stimulated by the presence of additional compounds which require reducing equivalen ts for further conversion, e.g. nitrite, or 2-oxoglutarate plus glutam ine. The [C-14]Glc6P-driven OPPP in isolated chromoplasts exhibited sa turation with rising concentrations of Glc6P, reaching highest rates a t an external concentration of about 2 mM. Exogenously given [(UC)-C-1 4]glucose 1-phosphate (Glc1P)' did not lead to a release of (CO2)-C-14 , indicating that this hexose phosphate is not taken up into the intac t plastid. Using a proteoliposome system in which the envelope membran e proteins from sweet-pepper chromoplasts were functionally reconstitu ted we demonstrated that Glc6P is transported in counter-exchange with inorganic phosphate (P-i) or other phosphorylated intermediates. The Glc6P was taken up into proteoliposomes with an apparent K-m of 0.34 m M. Surprisingly, in contrast to tomato fruit plastids, isolated chromo plasts from sweet-pepper fruits do not possess a phosphate translocato r allowing the uptake of Glc1P. Rising exogenous concentrations of dih ydroxyacetone phosphate strongly inhibited the metabolic flux through the OPPP. This observation is discussed with respect to the presence o f two phosphate translocator proteins in the envelope of sweet-pepper chromoplasts and with respect to possible metabolic changes occurring in heterotrophic tissues during development.