Amylases and Branching Enzyme of Developing Kidney Bean Seeds on Native Electrophoretic Gel

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  • トラマメ登熟種子中におけるアミラーゼおよび枝つけ酵素
  • Amylases and Branching Enzyme of Develo

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Abstract

When the crude extract from developing kidney bean (Phaseolus vulgaris L.) seeds was incubated soluble starch, β-limit dextrin or maltose, hydrolyzing activities were detected. The activity staining of gels of native polyacrylamide gel electrophoresis (i.e., zymograms) at pH 5.6 showed three bands on soluble starch and two on β-limit dextrin. Two active bands on both substrates were to heat treatment (at 70°C), while the other band from soluble starch had no activity. These results suggest that the heat-stable active bands correspond to α-amylases and the other to β-amylasea branching enzyme. Western blot analysis revealed that some bands bound to antiserum against amylase purified from germinating seeds. The zymogram at pH 7.0 indicated additional active from those observed at pH 5.6 on amylose. The mobilities of these two newly detected bands coincident with branching enzymes 1 and 2 purified from developing kidney bean seeds, suggesting the active band observed on soluble starch but not on β-limit dextrin at pH 5.6 was β-amylase. When the heat-treated crude extract was incubated with fl-limit dextrin at pH 5.6, glucose, maltose, other oligosaccharides were produced as evidenced by thin-layer chromatography, similar to α-amylase purified from germinating seeds. These data indicate that α-amylase exists in developing seeds.

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