Metabolite pools during starch synthesis and carbohydrate oxidation in amyloplasts isolated from wheat endosperm

Abstract

Starch synthesis and CO2 evolution were determined after incubating intact and lysed wheat (Triticum aestivum L. cv. Axona) endosperm amyloplasts with 14C-labelled hexose-phosphates. Amyloplasts converted [U-14C]glucose 1-phosphate (Glc1P) but not [U-14C]glucose 6-phosphate (Glc6P) into starch in the presence of ATP. When the oxidative pentose-phosphate pathway (OPPP) was stimulated, both [U-14C]Glc1P and [U-14C]Glc6P were metabolized to CO2, but Glc6P was the better precursor for the OPPP, and Glc1P-mediated starch synthesis was reduced by 75%. In order to understand the basis for the partitioning of carbon between the two potentially competing metabolic pathways, metabolite pools were measured in purified amyloplasts under conditions which promote both starch synthesis and carbohydrate oxidation via the OPPP. Amyloplasts incubated with Glc1P or Glc6P alone showed little or no interconversion of these hexose-phosphates inside the organelle. When amyloplasts were synthesizing starch, the stromal concentrations of Glc1P and ADP-glucose were high. By contrast, when flux through the OPPP was highest, Glc1P and ADP-glucose inside the organelle were undetectable, and there was an increase in metabolites involved in carbohydrate oxidation. Measurements of the plastidial hexose-monophosphate pool during starch synthesis and carbohydrate oxidation indicate that the phosphoglucose isomerase reaction is at equilibrium whereas the reaction catalysed by phosphoglucomutase is significantly displaced from equilibrium.