Glucose- and ADPGlc-dependent starch synthesis in isolated cauliflower-bud amyloplasts. Analysis of the interaction of various potential precursors.

Recently, we have demonstrated that isolated cauliflower-bud amyloplasts incorporate glucose 6-phosphate at high rates into newly synthesized starch (Neuhaus et al. (1993) Plant Physiol. 101, 573-578). Here we have analyzed the incorporation of radioactively labeled glucose and ADPglucose into newly synthesized starch. It could be shown that glucose incorporation into starch exhibits a typical substrate saturation behaviour and is linear with time for at least 40 min. The incorporation of glucose is strongly dependent upon the intactness of the plastids and upon the presence of both, ATP and 3-phosphoglyceric acid. Using 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) we showed that glucose is taken up into isolated cauliflower-bud amyloplasts as the free glucose molecule, rather than as glucose 6-phosphate. Glucose incorporation into newly synthesized starch is strongly inhibited in the presence of low concentrations of glucose 6-phosphate. The radioactively labeled glucose moiety of ADPglucose is also incorporated into starch. This incorporation can be saturated at increased concentrations of ADPglucose. ATP significantly inhibits the incorporation of the glucose moiety of ADPglucose into starch. This inhibition can be reinforced by the additional presence of glucose 6-phosphate. Glucose 6-phosphate-dependent starch synthesis is not strongly inhibited in the presence of glucose or ADPglucose indicating that glucose 6-phosphate is the precursor for starch synthesis in isolated cauliflower-bud amyloplasts.