Abstract
The enzyme ADP‐glucose pyrophosphorylase is essential for starch biosynthesis and is highly regulated. Here, mutations that increased heat stability and interactions with allosteric effectors were incorporated into the small subunit of the isoform known to be expressed at high levels in the maize endosperm. The resulting variants were transformed into maize with expression targeted to the endosperm. Transgenes harboring the changes increased yield some 35%; however, yield enhancement occurred via an increase in seed number rather than by increased seed weight. Interestingly, seed number increase is controlled by the genotype of the plant rather than the genotype of the seed as seeds increase in number whether or not they contain the transgene as long as the maternal parent has the transgene. The transgene is however expressed in the endosperm, and the altered allosteric and stability properties initially seen in Escherichia coli expression experiments are also seen with the endosperm‐expressed gene. The extent of seed number increase is positively correlated with the average daily high temperature during the first 4 days postpollination. While these results were unexpected, they echo the phenotypic changes caused by the insertion of an altered large subunit of this enzyme reported previously (Plant Cell, 24, 2012, 2352). These results call into question some of the reported fundamental differences separating starch synthesis in the endosperm vis‐à‐vis other plant tissues.
Highlights
Starch is the major component of cereal seeds and a critical constituent of cereal seed yields
A series of studies have pointed to the enzyme, ADP-glucose pyrophosphorylase (AGPase) as a key step in this important pathway
We showed that a chimeric small subunit in which the N-terminal 198 amino acids were derived from the BT2 protein and the carboxyl-terminal 277 amino acids originated from the potato tuber small subunit polymerized with the SH2 protein to produce an enzyme in an Escherichia coli expression system that exhibited a kcat some 10-fold greater than the wildtype maize endosperm in the absence of the activator, 3-phosphoglyceric acid (3PGA)
Summary
Starch is the major component of cereal seeds and a critical constituent of cereal seed yields. Placement of the cysteine-containing motif QTCL into the N-terminus of the chimeric MP small subunit enhanced heat stability more than 300-fold at 58°C in comparison with the wildtype maize endosperm enzyme (Boehlein et al, 2005; Linebarger et al, 2005). This alteration doubled the kcat of the enzyme, enhanced sensitivity to the activator 3PGA but did not affect Kms for the physiological substrates, ATP and glucose-1-phosphate or the Ki of phosphate. The unexpected finding of function in maternal tissue to increase seed number in a temperature-dependent manner has been found with an altered large subunit of the maize endosperm AGPase (Hannah et al, 2012)
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