title: Characterization of an autonomously activated plant ADP-glucose pyrophosphorylase.
abstract: ADP-glucose pyrophosphorylase (AGPase) catalyzes the rate-limiting step in starch biosynthesis in plants and changes in its catalytic and/or allosteric properties can lead to increased starch production. Recently, a maize (Zea mays)/potato (Solanum tuberosum) small subunit mosaic, MP [Mos(1-198)], containing the first 198 amino acids of the small subunit of the maize endosperm enzyme and the last 277 amino acids from the potato tuber enzyme, was expressed with the maize endosperm large subunit and was reported to have favorable kinetic and allosteric properties. Here, we show that this mosaic, in the absence of activator, performs like a wild-type AGPase that is partially activated with 3-phosphoglyceric acid (3-PGA). In the presence of 3-PGA, enzyme properties of Mos(1-198)/SH2 are quite similar to those of the wild-type maize enzyme. In the absence of 3-PGA, however, the mosaic enzyme exhibits greater activity, higher affinity for the substrates, and partial inactivation by inorganic phosphate. The Mos(1-198)/SH2 enzyme is also more stable to heat inactivation. The different properties of this protein were mapped using various mosaics containing smaller portions of the potato small subunit. Enhanced heat stability of Mos(1-198) was shown to originate from five potato-derived amino acids between 322 and 377. These amino acids were shown previously to be important in small subunit/large subunit interactions. These five potato-derived amino acids plus other potato-derived amino acids distributed throughout the carboxyl-terminal portion of the protein are required for the enhanced catalytic and allosteric properties exhibited by Mos(1-198)/SH2.
authors: Program in Plant Molecular and Cellular Biology and Horticultural Sciences, University of Florida, Gainesville, Florida 32611-7200, USA.
authors: Susan K
authors: SK
authors: Boehlein
authors: Janine R
authors: JR
authors: Shaw
authors: Jon D
authors: JD
authors: Stewart
authors: L Curtis
authors: LC
authors: Hannah
mesh_terms: D020816:Amino Acid Motifs; D004795:Enzyme Stability; D018506:Gene Expression Regulation, Plant; D051599:Glucose-1-Phosphate Adenylyltransferase / Q000235:genetics / Q000378:metabolism*; D005988:Glyceric Acids / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D011198:Solanum tuberosum / Q000201:enzymology* / Q000235:genetics; D003313:Zea mays / Q000201:enzymology* / Q000235:genetics
chemical_list: D005988:Glyceric Acids; D011994:Recombinant Proteins; C005156:3-phosphoglycerate; D051599:Glucose-1-Phosphate Adenylyltransferase