麹菌(Aspergillus oryzae)のグルコアミラーゼ遺伝子に関する研究

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Aspergillus oryzae, the most commonly used fungus in the Japanese fermentation industry, is of considerable industrial significance due to its ability to secrete copious quantities of hydrolytic enzymes. Some hydrolytic enzymes-especially glucoamylases, proteinases, and phosphatases-important in fermentation processes are produced in much higher amounts in solid-state culture than in submerged culture. However, the mechanism of the solid-state culture of this organism, including its higher enzyme productivity, remains unclear. Here, we describe the cloning and characterization of two glucoamylase-encoding genes, glaA and glaB, and examine their expression in order to clarify the mechanism of higher glucoamylase production in solid-state culture of A. oryzae. Sequence analysis of the two genes revealed that the glucoamylase encoded by glaA consists of two functional domains, a catalytic domain at the N-terminus and a starch-binding domain at the C-terminus, as is the case other Aspergillus glucoamylases. However, the glucoamylase encoded by glaB has no C-terminal domain related to raw starch binding activity. The glaB gene is markedly expressed in solid-state culture, but only a little in submerged culture. To elucidate the mechanism of glaB induction in solid-state culture, we analyzed the promoter using the sidA gene (GUS gene) as a reporter gene. In solid-state culture, glaB expression at the transcriptional level is enhanced by low-Aw (water activity), high-temperature, and physical barriers to hyphal extension, as well as by starch. Deletion analysis of the promoter showed that substitution of the 12 bp GC-rich motif from-335 to-324 (GC box) resulted in significant loss of starch and low-Aw induction. These findings suggest that the GC box is a cis-element essential for the high-level expression of glaB in solid-state culture.
社団法人日本生物工学会の論文
2000-04-25

麹菌(Aspergillus oryzae)のグルコアミラーゼ遺伝子に関する研究

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