Kim Sunhwa, Matsuo Ichiro, Ajisaka Katsumi, Nakajima Harushi, Kitamoto Katsuhiko
Department of Biotechnology, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
Biosci Biotechnol Biochem. 2002 Oct;66(10):2168-75. doi: 10.1271/bbb.66.2168.
We isolated a beta-N-acetylglucosaminidase encoding gene and its cDNA from the filamentous fungus Aspergillus nidulans, and designated it nagA. The nagA gene contained no intron and encoded a polypeptide of 603 amino acids with a putative 19-amino acid signal sequence. The deduced amino acid sequence was very similar to the sequence of Candida albicans Hex1 and Trichoderma harzianum Nag1. Yeast cells containing the nagA cDNA under the control of the GAL1 promoter expressed beta-N-acetylglucosaminidase activity. The chromosomal nagA gene of A. nidulans was disrupted by replacement with the argB marker gene. The disruptant strains expressed low levels of beta-N-acetylglucosaminidase activity and showed poor growth on a medium containing chitobiose as a carbon source. Aspergillus oryzae strain carrying the nagA gene under the control of the improved glaA promoter produced large amounts of beta-N-acetylglucosaminidase in a wheat bran solid culture.
我们从丝状真菌构巢曲霉中分离出一个编码β-N-乙酰氨基葡萄糖苷酶的基因及其cDNA，并将其命名为nagA。nagA基因不含内含子，编码一个由603个氨基酸组成的多肽，带有一个推定的19个氨基酸的信号序列。推导的氨基酸序列与白色念珠菌Hex1和哈茨木霉Nag1的序列非常相似。在GAL1启动子控制下含有nagA cDNA的酵母细胞表达β-N-乙酰氨基葡萄糖苷酶活性。用argB标记基因替换构巢曲霉的染色体nagA基因，使其发生破坏。破坏菌株表达的β-N-乙酰氨基葡萄糖苷酶活性水平较低，并且在以壳二糖作为碳源的培养基上生长不良。在改良的glaA启动子控制下携带nagA基因的米曲霉菌株在麦麸固体培养物中产生大量的β-N-乙酰氨基葡萄糖苷酶。
