Starch synthesis in the cereal endosperm
Introduction
Cereal crops accumulate starch in the seed endosperm as an energy reserve. This starch serves as the primary carbohydrate component in the diets of humans and livestock, and also has numerous important industrial applications. Starch comprises two d-glucose homopolymers, amylose and amylopectin. Amylose is essentially a linear molecule, in which glucosyl monomers are joined via α-1,4 linkages. Amylopectin, the more abundant polymer in starch, contains linear chains of various lengths. Approximately 5% of the glucosyl units in amylopectin are joined via α-1,6 linkages, which introduce chain branches. Amylopectin has a high degree of structural organization, as exemplified by the non-random distribution of linear chains and the clustered positioning of branch linkages. Regions of high-branch frequency alternate with regions that are devoid of branches, enabling intervening linear chains to align in parallel arrays of double helices (Figure 1; 1., 2.). This conserved architecture is responsible for the semi-crystalline nature of starch granules, which allows the dense packaging of glucose units. A higher-order organization in amylopectin gives rise to two types of crystalline structure, A-type and B-type, which differ with respect to the symmetry and packing of short amylopectin chains 3., 4.. Wildtype cereal starches are 100% A-type, in which double helices are arranged with a minimal amount of bound water.
This review covers recent advances in understanding the enzymatic activities necessary for starch synthesis and the determination of amylopectin structure, focusing particularly on new information on ADP-glucose pyrophosphorylase (AGP), starch synthase (SS), starch branching enzyme (BE) and starch debranching enzyme (DBE) 1., 2., 5.•. Biochemical characterizations, expression analyses, and genetic and transgenic approaches have combined to provide insight into the roles of specific enzyme isoforms and factors that potentially regulate their activities. Furthermore, owing to the sequencing of the rice genome, genome-based approaches for the examination of the starch biosynthetic pathway in cereals are now available for the first time.
Section snippets
AGP is uniquely extra-plastial in the cereal endosperm
AGP catalyzes the first reaction in starch synthesis, producing the activated glucosyl donor ADP-glucose (ADPG). AGP comprises two large subunits and two small subunits, each of which is encoded by distinct genes. The enzyme is now known to be largely extra-plastidial (i.e. 85–95% cytosolic) in cereal endosperm, but plastidial in other cereal tissues and in all tissues of non-cereal plants (Figure 2; 6., 7., 8., 9., 10.••, 11.). Recent studies suggest that distinct cytosolic and plastidial
Individual starch synthase isoforms have unique roles in starch synthesis
SSs utilize ADPG to elongate linear chains by the formation of α-1,4 linkages. Cereal endosperms contain at least five SS isoforms that are categorized according to conserved sequence relationships [23]. Four isoforms, termed SSI, SSIIa, SSIIb, and SSIII, are believed to have unique functions in amylopectin synthesis, although their precise roles have not been identified. A granule-bound isoform, GBSSI, which is encoded by the Waxy (Wx) locus in cereals, functions specifically to elongate
Contributions of individual BE isoforms to the determination of starch structure
BEs generate α-1,6 linkages by cleaving internal α-1,4 bonds and transferring the released reducing ends to C6 hydroxyls. There are two classes of BE (BEI and BEII) that differ in terms of the lengths of chains transferred in vitro, with BEII transferring shorter chains than BEI 35., 36.. In cereals, there are two closely related forms of BEII (BEIIa and BEIIb) 37.•, 38.. These also differ in chain-length specificity in vitro, with BEIIb transferring shorter chains than BEIIa during extended
Potential DBE functions
Mutations in many species indicate that starch synthesis involves DBEs in addition to SSs and BEs. Two DBE families exist in plants, isoamylase-type and pullulanase-type. Both types hydrolyze α-1,6 linkages, but they differ in substrate specificity. Orthologous mutations of maize and rice (sugary1 [su1]) and barley (isa-1) affect genes that encode an isoamylase-type DBE, and correlate with the accumulation of a polymeric water-soluble polysaccharide (WSP) termed phytoglycogen and reduced starch
Protein modifications that potentially regulate starch biosynthesis
Little is known about factors that regulate starch biosynthesis in cereals. Recent research suggests that some regulation occurs through protein modifications, which are also implicated in the control of other pathways. Reduction of the activity of SPK, a calcium-dependent protein kinase, in developing rice endosperm reduced the starch content and increased the sucrose content of the grain [53•]. SPK phosphorylates sucrose synthase, the enzyme that provides substrate for AGP, and thus may
Conclusions and future directions
Our understanding of starch synthesis in cereal endosperms has advanced recently as we have gained insights into the specific functions of individual enzyme isoforms. A cytosolic form of AGP that is unique to cereals may serve to commit excess carbon to starch production. Roles for SSI, SSIIa, and all three BEs in chain-length determination and branch placement have been proposed. The pleiotropic effects of BE and DBE mutants complicate the analyses, and have led to additional hypotheses
References and recommended reading
Papers of particular interest, published within the annual period of review, have been highlighted as:
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of special interest
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of outstanding interest
Acknowledgements
The authors thank Alison Smith for her helpful suggestions in the preparation of this article. We gratefully acknowledge the US Department of Agriculture, the US Department of Energy, and the National Science Foundation for providing research funding.
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