REGULATION OF MAIZE LEAF SUCROSE-PHOSPHATE SYNTHASE BY PROTEIN-PHOSPHORYLATION

Studies were conducted to determine the potential for regulation of maize leaf sucrose-phosphate synthase (SPS) by protein phosphorylation. Highly activated enzyme, in desalted crude leaf extracts prepared from illuminated leaves, was inactivated in vitro in a time- and ATP-dependent manner. Partial purification of SPS by polyethylene glycol fractionation and Mono Q chromatography yielded enzyme that was not ATP-inactivated, possibly due to elimination of contaminating protein kinase. We used the partially purified SPS as substrate to identify an endogenous protein kinase. The protein kinase catalyzed the time- and ATP-dependent inactivation of SPS, and the apparent K(m) for Mg.ATP was estimated to be approximately 10-mu-M. The partially purified maize SPS protein was phosphorylated in vitro using [gamma-P-32]ATP and either the endogenous protein kinase or the catalytic subunit of cAMP-dependent protein kinase. The incorporation of radiolabel was closely paralleled by inactivation of the enzyme. These results provide the first evidence for regulation of maize leaf SPS by protein phosphorylation, which we postulate is the mechanism of light-dark regulation in vivo.