Abstract
It is known that one of starch branching enzyme (BE) isoforms, BEIIb, plays a specific role not only in the synthesis of distinct amylopectin cluster structure, but also in the formation of the internal structure of starch granules in rice endosperm because in its absence the starch crystalline polymorph changes to the B-type from the typical A-type found in the wild-type (WT) cereal endosperm starch granules. In the present study, to examine the contribution of BEIIb to the amylopectin cluster structure, the chain-length distributions of amylopectin and its phosphorylase-limit dextrins (Φ-LD) from endosperm and culm of a null be2b mutant called amylose-extender (ae) mutant line, EM10, were compared with those of its WT cultivar, Kinmaze, of japonica rice. The results strongly suggest that BEIIb specifically formed new short chains whose branch points were localized in the basal part of the crystalline lamellae and presumably in the intermediate between the crystalline and amorphous lamellae of amylopectin clusters in the WT endosperm, whereas in its absence branch points which were mainly formed by BEI were only located in the amorphous lamellae of amylopectin. These differences in the cluster structure of amylopectin between Kinmaze and EM10 endosperm were considered to be responsible for the differences in the A-type and B-type crystalline structures of starch granules between Kinmaze and EM10, respectively. The changes in internal structure of starch granules caused by BEIIb were analyzed by wide angle X-ray diffraction, small-angle X-ray scattering, solid state 13C NMR, and optical sum frequency generation spectroscopy. It was noted that the size the amylopectin cluster in ae endosperm (approximately 8.24 nm) was significantly smaller than that in WT endosperm (approximately 8.81 nm). Based on the present results, we proposed a model for the cluster structure of amylopectin in WT and ae mutant of rice endosperm. We also hypothesized the role of BEIIa in amylopectin biosynthesis in culm where BEIIb was not expressed and instead BEIIa was the major BE component in WT of rice.
Highlights
Starch is composed of an essentially linear or scarcely branched glucan called amylose and a highly branched glucan called amylopectin, both components accounting for 15–30% and 70– 85%, respectively
Since BEIIb is expressed in endosperm of cereals such as maize and rice, but not in tubers, this isozyme must play a critical role in the formation of the cereal-specific amylopectin cluster structure
These changes in the cluster structure of amylopectin induced by the be2b mutation caused the internal structure of starch granules from the A-type to B-type polymorph, as detected by X-ray diffraction (XRD), small angle X-ray scattering (SAXS), solid state 13C NMR, and Sum Frequency Generation (SFG) analyses
Summary
Starch is composed of an essentially linear or scarcely branched glucan called amylose and a highly branched glucan called amylopectin, both components accounting for 15–30% and 70– 85%, respectively. Amylopectin has a unit structure designated as cluster whereas glycogen has no such unit structure because upalpha-1,6 branch points are randomly distributed in glycogen (Thompson, 2000). The clusters of amylopectin are interconnected by long chains designated as B2 chains and/or B3 chains that span to two clusters and/or three clusters, respectively, whereas short and intermediate chains within a single cluster are called A chains (non-branched chains) or B1 chains (branched by at least one chains) (Peat et al, 1952). After the treatment of debranching with isoamylase the chain-length distribution of amylopectin forms a bimodal pattern whereas that of glycogen shows only a single peak in addition to the longer average chain-length in amylopectin compared with glycogen
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