Biosynthesis of Starch: ADPglucose Pyrophosphorylase, the Regulatory Enzyme of Starch Synthesis: Structure-Function Relationships.

The synthesis of starch is believed to involve 3 enzymes, ADPglucose pyrophosphorylase (ADPGIc PPase, [EC 2.7.7.27] ; reaction 1), starch synthase, (EC 2.4.1.21) ; reaction 2) and branching enzyme ([EC 2.4.1.18] ; reaction 3). (1) α-glucosyl-l-P + ATP ↔ ADPGIc + PPi (2) ADPGIc + (glucosyl) n → ADP + (glucosyl) n+1 (3) Linear α-1, 4-glucosyl chain of α-glucan→Amylopectin (a-1, 6 and 1, 4-glucan) There are many genetic studies to indicate that all three reactions contribute to the synthesis of both starch components, amylose and amylopectin. Maize amylose extender (ae) mutants, containing more amylose and less amylopectin than normal maize have been shown to be defective in one of the branching enzyme isoforms (Plant Physiol., 67, 1141-1145 (1981)). Maize waxy (wx) mutants which contain no amylose are deficient in the starch granule bound starch synthase (Plant Physiol., 62, 383-386 (1978)) indicating that the granule bound starch synthase is involved in amylose synthesis. Starch deficient mutants of pea (Plant Physiol., 89, 1279-1284 (1989)), maize (Science, 151, 343-348 (1966)) and Arabidopsis (Plant Physiol., 86, 1131-1135 (1988)) are deficient in ADPGIc PPase activity. Thus, genetic and biochemical evidence in plants strongly indicate that ADPglucose (ADPGIc) is synthesized predominantly, if not solely, via ADPGIc PPase catalysis. The cyanobacterial, algal and higher plant enzymes are activated by 3-P-glycerate (3PGA) and inhibited by orthophosphate. An algal mutant in which the regulatory sites of the ADPGIc PPase have been affected (Planta, 185, 17-26) (1991) and control analysis (Kacsar-Burns) of the plant ADPGIc PPase, indicate that the allosteric regulation observed of the enzyme in plants reflects the in vivo regulation of plant starch synthesis. Thus, the above studies empha-size the importance of the ADPGIc PPase in starch synthesis and the need for understanding the structure-function relationships of the regulatory and catalytic domains of the ADPGIc PPase. A number of chemical modification and site-directed mutageneses studies have identified the active and substrate binding sites both in the spinach leaf and bacterial enzymes. There is considerable conservation between the bacterial and higher plant enzymes particularly at the substrate binding sites. High conservation is also observed for the 3PGA activator sites of the cyanobacterial and higher plant ADPGIc PPases. These results suggest that the two subunits of the higher plant ADPGIc PPase evolved from the bacterial enzyme subunit. Transformation of the plant systems with the bacterial enzyme gene results in an appreciable increase in starch synthesis ranging from 1.25-to 1.6-fold in potato tubers to 10-fold in tobacco calli. These results confirm that the ADPGIc PPase activity in plants is rate-limiting for starch synthesis and that the allosteric regulation is functional in the plant.