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Enzymic Mechanism of Starch Breakdown in Germinating Rice Seeds

10. IN VIVO AND IN VITRO SYNTHESIS OF -AMYLASE IN RICE SEED SCUTELLUM

Research Institute for Biochemical Regulation, School of Agriculture, Nagoya University, Chikusa, Nagoya 464, Japan

Scutellar tissues were dissected from germinating rice seeds and the incorporation of [³⁵S]methionine into the -amylase molecule was examined by in vivo and in vitro assay systems. Immunoprecipitation with monospecific anti--amylase immunoglobulin G raised against the purified enzyme preparation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography were used to identify -amylase and its possible precursor molecule. Using freshly prepared scutellar tissues, it was demonstrated that -amylase is synthesized de novo in the scutellar epithelium and secreted into endosperm. The synthesis of -amylase directed by the polyadenylic acid-containing ribonucleic acid isolated from the scutellar tissues was also established using the translation system of either wheat germ extract or reticulocyte lysate. The immunoprecipitable product obtained in the in vitro translation system was smaller in molecular weight than that synthesized in vivo on the basis of mobilities in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Results are discussed in relation to the processing of the nascent polypeptide precursor of the enzyme molecule and the introduction of the oligosaccharide chain to the cleaved polypeptide to make up the mature form of -amylase.

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