Abstract
Starch branching enzyme (SBE, EC 2.4.1.18) activity was followed in developing barley (Hordeum vulgare L. cv. Golf) caryopses during a period of one month after anthesis. Caryopses with the highest specific activity, and corresponding to a fresh weight of around 60 mg per caryopsis, were homogenized and the soluble extract used for branching enzyme purification by FPLC chromatography. Four branching enzyme activity fractions were resolved. From one of these fractions, which exhibited high activity in both the phosphorylation stimulation and amylose branching assays, a branching enzyme preparation containing two related polypeptides of 51 and 50 kDa was obtained. Native polyacrylamide gel electrophoresis and gel filtration showed that the 51/50‐kDa polypeptide is monomeric. A combination of phosphorylation stimulation and amylose branching gel assays, SDS‐PAGE, and TLC was used to demonstrate the branching activity of the 51/50‐kDa polypeptide. The activity was further confirmed by spectroscopic analyses of iodine‐glucan complexes. SBEs from four different plant species were compared using the phosphorylation stimulation gel assay.
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