Regulatory mechanisms of xpression of glycosidase genes in filamentous fungi and their applications

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Filamentous fungi represented by Aspergillus oryzae have been playd important roles in Japanese fermentation industries. We have examined the expression properties of Taka-amylase A gene isolated from A.oryzae using A.nidulans as an intermediate host. CCAAT sequence and AnCP,CCAAT binding protein, have been shown to play important roles in enhancement of Taka-amylase A gene expression under the inducible conditions.In this project, cellulase and xylanase genes have been characterized to elucidate their regulatory mechanisms in addition to Taka-amylase gene in order to develop a novel way to utilize efficiently various reproductive natural resources, especially carbohydrates such as starch, cellulose and xylan. We have obtained the following results during last two years.1. HAPC,a compoment of AnCP,was prepared by expressing malE-hapC fusion gene in E.coli and used to raise an antibody to HAPC.With recombinant HAPC and its antibody, we succedeed to prove that AnCP comprised three subunits including HAPC.2. CCAAT sequence was proved not to be involved in the induction of the gene. SRE (starch responsive element) and its binding protein, SREB,were suggested to play a vital role in induction of Taka-amylaseA gene.3. CMC inducible cellulase gene, eglA,was isolated from A.nidulans and sequenced. Preliminary analysis as to the promoter region was also conducted to find any elements for induction or reperssion of eglA.4. Xylanase gene, cgxA,isolated from C.gracile was characterized for itsrepressive element and repressor-likefactor which recognized the 13bp repressive element. Furthermore, the A.oryzae xylanase gene, xynFI,was also determined for its inducible element. A 35bp element appeared to be involved in inductive expression of the gene.