title: Three extracellular dipeptidyl peptidases found in Aspergillus oryzae show varying substrate specificities.
abstract: Three extracellular dipeptidyl peptidase genes, dppB, dppE, and dppF, were unveiled by sequence analysis of the Aspergillus oryzae genome. We investigated their differential enzymatic profiles, in order to gain an understanding of the diversity of these genes. The three dipeptidyl peptidases were expressed using Aspergillus nidulans as the host. Each recombinant enzyme was purified and subsequently characterized. The enzymes displayed similar optimum pH values, but optimum temperatures, pH stabilities, and substrate specificities varied. DppB was identified as a Xaa-Prolyl dipeptidyl peptidase, while DppE scissile substrates were similar to the substrates for Aspergillus fumigatus DPPV (AfDPPV). DppF was found to be a novel enzyme that could digest both substrates for A. fumigatus DPPIV and AfDPPV. Semi-quantitative PCR revealed that the transcription of dppB in A. oryzae was induced by protein substrates and repressed by the addition of an inorganic nitrogen source, despite the presence of protein substrates. The transcription of dppE depended on its growth time, while the transcription of dppF was not affected by the type of the nitrogen source in the medium, and it started during the early stage of the fungal growth. Based on these results, we conclude that these enzymes may represent the nutrition acquisition enzymes. Additionally, DppF may be one of the sensor peptidases responsible for the detection of the protein substrates in A. oryzae environment. DppB may be involved in nitrogen assimilation control, since the transcription of dppB was repressed by NaNO3, despite the presence of protein substrates.
authors: Department of Applied Molecular Biology and Biochemistry, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu, Tokyo, 1838509, Japan.
authors: Hiroshi
authors: H
authors: Maeda
authors: Department of Applied Molecular Biology and Biochemistry, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu, Tokyo, 1838509, Japan.
authors: Daisuke
authors: D
authors: Sakai
authors: Department of Applied Molecular Biology and Biochemistry, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu, Tokyo, 1838509, Japan.
authors: Takuji
authors: T
authors: Kobayashi
authors: Department of Applied Molecular Biology and Biochemistry, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu, Tokyo, 1838509, Japan.|Microbiology & Fermentation Laboratory, CALPIS Co. Ltd. 5-11-10 Fuchinobe, Chuo-ku, Sagamihara, Kanagawa, 2520206, Japan.
authors: Hiroto
authors: H
authors: Morita
authors: Department of Applied Molecular Biology and Biochemistry, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu, Tokyo, 1838509, Japan.
authors: Ayako
authors: A
authors: Okamoto
authors: Department of Applied Molecular Biology and Biochemistry, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu, Tokyo, 1838509, Japan.
authors: Michio
authors: M
authors: Takeuchi
authors: National Food Research Institute, 2-1-12 Kan-nondai, Tsukuba, Ibaraki, 3058642, Japan.
authors: Ken-Ichi
authors: K
authors: Kusumoto
authors: Amano Enzyme Inc., 1-2-7 Nishiki, Naka-ku, Nagoya, Aichi, 4608630, Japan.
authors: Hitoshi
authors: H
authors: Amano
authors: Gekkeikan Sake Co., Ltd., 247 Minamihama-cho, Fushimi-ku, Kyoto, 6128660, Japan.
authors: Hiroki
authors: H
authors: Ishida
authors: Department of Applied Molecular Biology and Biochemistry, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu, Tokyo, 1838509, Japan. y-yama@cc.tuat.ac.jp.
authors: Youhei
authors: Y
authors: Yamagata
mesh_terms: D001232:Aspergillus fumigatus / Q000201:enzymology / Q000235:genetics; D001233:Aspergillus nidulans / Q000201:enzymology / Q000235:genetics; D001236:Aspergillus oryzae / Q000201:enzymology* / Q000235:genetics*; D004271:DNA, Fungal / Q000302:isolation & purification; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D004271:DNA, Fungal; D011994:Recombinant Proteins; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases
keywords: Aspergillus oryzae; Dipeptidyl peptidase; Diversity of proteolytic enzymes; Proteolytic enzyme; Substrate specificity