Maize Starch-Branching Enzyme Isoforms and Amylopectin Structure. In the Absence of Starch-Branching Enzyme IIb, the Further Absence of Starch-Branching Enzyme Ia Leads to Increased Branching

Abstract

Previous studies indicated that the deficiency of starch-branching enzyme (SBE) Ia in the single mutant sbe1a::Mu (sbe1a) has no impact on endosperm starch structure, whereas the deficiency of SBEIIb in the ae mutant is well known to reduce the branching of starch. We hypothesized that in maize (Zea mays) endosperm, the function of SBEIIb is predominant to that of SBEIa, and SBEIa would have an observable effect only on amylopectin structure in the absence of SBEIIb. To test this hypothesis, the mutant sbe1a was introgressed into lines containing either wx (lacking the granule-bound starch synthase GBSSI) or ae wx (lacking both SBEIIb and GBSSI) in the W64A background. Both western blotting and zymogram analysis confirmed the SBEIa deficiency in sbe1a wx and sbe1a ae wx, and the SBEIIb deficiency in ae wx and sbe1a ae wx. Using zymogram analysis, no pleiotropic effects of sbe1a genes on SBEIIa, starch synthase, or starch-debranching enzyme isoforms were observed. Highperformance size exclusion chromatography analysis shows that the chain-length profiles of amylopectin as well as b-limit dextrin were indistinguishable between wx and sbe1a wx, whereas significant differences for both were observed between ae wx and sbe1a ae wx, suggesting an effect of SBEIa on amylopectin biosynthesis that is observable only in the absence of SBEIIb. The amylopectin branch density and the average number of branches per cluster were both higher in endosperm starch from sbe1a ae wx than from ae wx. These results indicate possible functional interactions between SBE isoforms that may involve enzymatic inhibition. Both the cluster repeat distance and the distance between branch points on the short intracluster chains were similar for all genotypes however, suggesting a similar pattern of individual SBE isoforms in cluster initiation and the determination of branch point location.Starch biosynthesis involves the activities of four enzymes: ADP-Glc pyrophosphorylase, starch synthase (SS), starch-branching enzyme (SBE), and starch-debranching enzyme (DBE;Smith et al., 1997). The coordinated functions of these enzymes result in two forms of starch molecules: amylose, an essentially linear glucan chain consisting of a-1,4 linkages, and amylopectin, a highly branched glucan chain with multiple branch points formed by a-1,6 linkages.SBE catalyzes the formation of branch points within glucan chains by cleaving an a-1,4 linkage and reattaching the chain to a glucan chain via a-1,6 bond. Three isoforms of SBE (SBEIa, SBEIIa, and SBEIIb) have been identified in maize (Zea mays; Boyer and Preiss, 1978;Dang and Boyer, 1988), and their expression patterns differ considerably (Gao et al., 1996). While the in vitro properties of SBEIa differ from those of SBEIIa and SBEIIb (Guan and Preiss, 1993;Takeda et al., 1993), the sbe1a or sbe2a single mutants obtained using Mu-mediated transposon mutagenesis indicated that the chain-length (CL) profiles of starch produced in endosperm in vivo were not affected by the deficiency of either SBEIa or SBEIIa (Blau...

A review of starch‐branching enzymes and their role in amylopectin biosynthesis

“…Interestingly, loss of SBEI activity in monocots and dicots has minimal apparent effects on starch synthesis and composition (45), although in a study by Yao et al, loss of SBEI from a SBEIIbdeficient background in maize was shown to cause increased branching of amylopectin. Yao et al (46) postulated that this result was suggestive of a regulatory role for SBEI in influencing other SBE isoforms. Indeed, physical interactions between SBEI and SBEIIb have been reported in cereal endosperm amyloplasts (see later section).…”

Mutants of Arabidopsis Lacking Starch Branching Enzyme II Substitute Plastidial Starch Synthesis by Cytoplasmic Maltose Accumulation

“…Mutant lines defective for SBEI, IIa, and IIb in maize (Stinard et al, 1993;Blauth et al, 2001Blauth et al, , 2002Yao et al, 2004) and rice (Oryza sativa; Nishi et al, 2001;Nakamura, 2002;Satoh et al, 2003) were isolated and analyzed for their ability to synthesize starch. The absence of SBEI in both plants has little or no impact on starch that accumulates in the endosperm, while the absence of SBEIIb leads to the synthesis of a modified amylopectin with a strong decrease in the number of short chains and an increase in the number of long chains.…”

Recent progress toward understanding the role of starch biosynthetic enzymes in the cereal endosperm

“…An in vitro study showed that SBEI transfers longer chains (DP ≥ 16) and has a higher affinity for amylose, while SBEII transfers shorter chains (DP ≤ 12) and appears to have a higher affinity for amylopectin [45]. However, in maize endosperm the amylopectin CLD is not affected by a deficiency in SBEI activity, although a further loss of SBEIIb causes increased branching, suggestive of a regulatory role for SBEI in influencing other SBEs [77,78]. This is supported by the observation of the physical interaction between SBEI and SBEIIb in the wheat endosperm amyloplast [71].…”