Plant Physiology 98:1277-1284 (1992)
© 1992 American Society of Plant Biologists
Metabolism and Enzymology
Characterization and Subcellular Localization of Debranching Enzyme and Endoamylase from Leaves of Sugar Beet 1
Bin Li,
Jerome C. Servaites and
Donald R. Geiger
Department of Biology, University of Dayton, Dayton, Ohio 45469-2320
Sugar beet leaves (Beta vulgaris L.) contained up to five endoamylases, two exoamylases, and a single debranching enzyme. Four of the endoamylases and the debranching enzyme were present in the chloroplast. The chloroplastic starch-debranching enzyme and an apoplastic endoamylase were copurified from mature leaves of sugar beet by 35 to 50% ammonium sulfate precipitation and chromatography on diethylaminoethyl-Sephacryl, -cyclodextrin Sepharose 6B, and Sephadex G-150. The debranching enzyme, which was purified to homogeneity, had a molecular mass of 100 kilodaltons and a pH optimum of 5.5. It showed a high activity with pullulan as a substrate, low activity with soluble starch and amylopectin, and no activity with native starch grains isolated from sugar beet leaves. The endoamylase, which was partially purified, had a molecular mass of 43,000 kilodaltons, a pH optimum of 6.5, required calcium for activity and thermal stability, and showed an ability to hydrolyze native starch grains.
1 Supported by National Science Foundation grants DCB 88-16970 and 89-15789, the Ohio Board of Regents/Research Challenge Program, and summer fellowships from the University of Dayton Research Council.
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