Cloning, Sequencing, and Expression of a Gene Encoding the Monomeric Isocitrate Dehydrogenase of the Nitrogen-fixing Bacterium, Azotobacter vinelandii(Biochemistry & Molecular Biology)
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概要
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Isocitrate dehydrogenase (IDH : EC 1. 1. 1. 42) of Azotobacter vinelandii was purified to an electrophoretically homogeneous state, and a gene (icd) encoding this enzyme was cloned and sequenced. The N-terminal amino acid sequence of the purified enzyme was consistent with that deduced from the nucleotide sequence of the icd gene. The deduced amino acid sequence of this gene showed high identity (62-66%) to those of the other bacterial monomeric IDHs. Expression of the icd gene in Escherichia coli was examined by measuring the enzyme activity and mRNA level. Primer extension analyses revealed that two species of mRNAs with different lengths of 5'-untranslated regions (TS-1 and TS-2) were present, of which the 5'-terminals (TS-1 and TS-2 sites) were cytosines located at 244 bp and 101 bp upstream of translational initiation codon, respectively. Conserved promoter elements were present at -35 and -10 regions from the TS-1 site, whereas no such a common motif was found in the upstream region of the TS-2 site. Deletion of the promoter elements upstream of the TS-1 site resulted in complete loss of IDH activity in the E. coli transformant. When the promoter elements upstream of the TS-1 site were intact, the levels of TS-1 and TS-2 were varied greatly by altering exogenous nutrients for growth. The cells grown in a nutrient-rich medium produced large amounts of TS-1 and had a low level of IDH activity. In a nutrient-poor medium, the cells contained large amounts of TS-2 and high levels of IDH activity.
- 社団法人日本農芸化学会の論文
- 2002-03-23
著者
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Takada Yasuhiro
Division Of Biological Sciences Graduate School Of Science Hokkaido University
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Toba Y
Snow Brand Milk Products Co. Ltd. Saitama Jpn
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Toba Yasuhiro
Technology And Research Institute Snow Brand Milk Products Co. Ltd.
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Takada Yukihiro
雪印乳業技術研究所
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Takada Y
Snow Brand Milk Products Co. Ltd. Saitama Jpn
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Toba Y
Technology And Research Institute Snow Brand Milk Products Co. Ltd.
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Takada Y
Hokkaido Univ. Hokkaido Jpn
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Takada Yasuhiro
Hokkaido Univ. Sapporo Jpn
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FUKUNAGA NORIYUKI
Division of Biological Sciences, Graduate School of Science, Hokkaido University
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Takeuchi Yoji
Cell Function And Structure Division Of Biological Sciences Graduate School Of Science Hokkaido Univ
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SAHARA Takehiko
Division of Biological Sciences, Graduate School of Science, Hokkaido University
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TAKEUCHI Yoji
Division of Biological Sciences, Graduate School of Science, Hokkaido University
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YAMAOKA Naoto
Division of Biological Sciences, Graduate School of Science, Hokkaido University
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Fukunaga Noriyuki
Division Of Biological Sciences Graduate School Of Science Hokkaido University
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Takeda Yuichi
Fac. Sci. And Tech. Keio Univ. Yokohama
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Sahara Takehiko
Division Of Biological Sciences Graduate School Of Science Hokkaido University:(present Address)hokk
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Yamaoka N
Laboratory Of Plant Pathology Department Of Agriculture Ehime University
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