ADPglucose pyrophosphorylase (AGPase) catalyzes the first unique step in the starch biosynthetic pathway. Recent studies in barley, wheat, and maize indicate that the major endosperm enzyme exhibits different enzymatic and cellular properties than those found in leaf tissue. To determine how prevalent these properties are in other plants, the AGPase activities of rice endosperm were studied at the enzymatic and cellular levels. Unlike the barley and wheat endosperm enzymes, the rice AGPase was dependent on 3-phosphoglycerate (3-PGA) with more than 40-fold increase in catalytic activity, when assayed under near saturating 3-PGA conditions. The rice enzyme was also inhibited by Pi and this inhibition was reversed by 3-PGA. Subcellular fractionation studies indicated that the bulk (90%) of the AGPase activity was extra-plastidic, located in the cytoplasm. A smaller amount of enzyme activity (10%) was observed associated with the amyloplast fraction. The presence of two AGPase forms was supported by immunoblot analysis using anti-bodies specific for the large or small subunit of the heterotetrameric enzyme. Anti-bodies specific for the maize endosperm AGPase small subunit (BT2) recognized a major 54 kD polypeptide in the cytoplasmic fraction. A second polypeptide at 48 kD was also detected in the cytoplasmic fraction but was more abundant in the amyloplast fraction. Based on results obtained from this study, strategies for increasing starch synthesis by manipulation of AGPase activities in developing seeds and, in turn, seed yields are discussed.
Bibliographical noteFunding Information:
This study was supported in part by grants from the Department of Energy DEFG03-96ER20216 and US Department of Agriculture National Research Initiative Competitive Grants Program Award no. 98-35306-6469. V.S. was supported by a Rockefeller Foundation Program in Rice Biotechnology Fellowship. T.W.O. is a participant in the Hatch NC-142 Project. We thank Jack Preiss for the spinach leaf 54 kD anti-body, L. Curtis Hannah for the maize SH2 and BT2 anti-bodies and cDNA plasmids, and H. Satoh and T. Kumamaru for the rice developing seed cDNA library.
- ADPglucose pyrophosphorylase
- Oryza sativa